阅读说明：本技术 一种鉴定黄瓜种质真实性的snp位点引物组合及应用 (SNP (Single nucleotide polymorphism) site primer combination for identifying cucumber germplasm authenticity and application ) 是由 温常龙 杨静静 张建 罗江 刘庞源 毛爱军 王航 于 2020-04-20 设计创作，主要内容包括：本发明属于分子标记及其检测领域,具体涉及一种鉴定黄瓜种质真实性的核心SNP位点、引物组合,基于SNP位点的DNA指纹数据库、数据库构建方法,以及基于SNP位点的黄瓜种质真实性的鉴定方法、应用。所述SNP位点选自第一SNP位点到第四十八SNP位点中的任1到48种。本发明建立了基于高通量测序鉴定黄瓜种质真实性的DNA指纹数据库,可用于对黄瓜种质在种子、幼苗、叶片等组织或器官进行早期鉴定,为黄瓜种质资源保护提供技术支持。本发明提供的方法既可以对未知黄瓜种质进行鉴定,也可以对已知黄瓜种质的真实性进行鉴定。本发明提供的方法具有高通量、准确、低成本、操作简单、节约人力、物力等优点,具有十分广阔的应用前景。(The invention belongs to the field of molecular markers and detection thereof, and particularly relates to a core SNP (single nucleotide polymorphism) site and primer combination for identifying the authenticity of cucumber germplasm, a DNA (deoxyribonucleic acid) fingerprint database and database construction method based on the SNP site, and an identification method and application of the authenticity of cucumber germplasm based on the SNP site. The SNP sites are selected from any 1 to 48 of the first SNP site to the fourth eighteenth SNP site. The invention establishes a DNA fingerprint database for identifying the authenticity of the cucumber germplasm based on high-throughput sequencing, can be used for carrying out early identification on the cucumber germplasm in tissues or organs such as seeds, seedlings, leaves and the like, and provides technical support for cucumber germplasm resource protection. The method provided by the invention can be used for identifying unknown cucumber germplasm and also can be used for identifying the authenticity of the known cucumber germplasm. The method provided by the invention has the advantages of high throughput, accuracy, low cost, simplicity in operation, manpower and material resource saving and the like, and has a very wide application prospect.)

1. Core SNP loci for identifying the germplasm authenticity of cucumber, wherein the SNP loci are selected from any 1 to 48 of the following first SNP loci to fourth eighteen SNP loci:

a first SNP locus, wherein the first SNP locus is located at 215528 th chromosome 1 of a cucumber reference genome, and the nucleotide base of the first SNP locus is G or A;

a second SNP locus, wherein the second SNP locus is located at 6591765 th chromosome 1 of a cucumber reference genome, and the nucleotide base of the second SNP locus is A or T;

a third SNP locus, wherein the third SNP locus is located at 18360230 th chromosome 1 of a cucumber reference genome, and the nucleotide base of the locus is G or C;

a fourth SNP locus, wherein the fourth SNP locus is located at 19328118 th chromosome 1 of a cucumber reference genome, and the nucleotide base of the fourth SNP locus is T or C;

a fifth SNP locus, wherein the fifth SNP locus is located at 21471767 th chromosome 1 of a cucumber reference genome, and the nucleotide base of the locus is G or C;

a sixth SNP locus, wherein the sixth SNP locus is located at 26006098 th chromosome 1 of a cucumber reference genome, and the nucleotide base of the locus is A or G;

a seventh SNP locus, wherein the seventh SNP locus is located at 430448 th chromosome 2 of a cucumber reference genome, and the nucleotide base of the locus is C or T;

an eighth SNP locus, wherein the eighth SNP locus is located at 6544548 th chromosome 2 of a cucumber reference genome, and the nucleotide base of the locus is A or T;

a ninth SNP locus, wherein the ninth SNP locus is located at 7082492 th chromosome 2 of a cucumber reference genome, and the nucleotide base of the locus is A or T;

a tenth SNP locus, wherein the tenth SNP locus is located at 13016733 th chromosome 2 of a cucumber reference genome, and the nucleotide base of the locus is C or T;

an eleventh SNP locus, wherein the eleventh SNP locus is located at 19736406 th chromosome 2 of a cucumber reference genome, and the nucleotide base of the locus is C or T;

a twelfth SNP locus, wherein the twelfth SNP locus is located at 22890765 th chromosome 2 of the cucumber reference genome, and the nucleotide base of the locus is C or T;

a thirteenth SNP locus, wherein the thirteenth SNP locus is located at 3674759 th chromosome 3 of a cucumber reference genome, and the nucleotide base of the locus is G or A;

a fourteenth SNP locus, wherein the fourteenth SNP locus is located at 6715343 th chromosome 3 of a cucumber reference genome, and the nucleotide base of the locus is T or C;

a fifteenth SNP locus, wherein the fifteenth SNP locus is located at 8251132 th chromosome 3 of a cucumber reference genome, and the nucleotide base of the locus is G or A;

a sixteenth SNP locus, wherein the sixteenth SNP locus is located at 17274568 th chromosome 3 of a cucumber reference genome, and the nucleotide base of the sixteenth SNP locus is G or A;

a seventeenth SNP locus, wherein the seventeenth SNP locus is positioned at 28481554 rd chromosome of the cucumber reference genome, and the nucleotide base of the locus is C or T;

an eighteenth SNP locus, wherein the eighteenth SNP locus is located at 30072485 th chromosome of a cucumber reference genome 3, and the nucleotide base of the locus is A or G;

a nineteenth SNP locus, wherein the nineteenth SNP locus is located at 31558451 th chromosome 3 of a cucumber reference genome, and the nucleotide base of the locus is A or G;

a twentieth SNP locus, wherein the twentieth SNP locus is located at 1284842 th chromosome 4 of the cucumber reference genome, and the nucleotide base of the locus is G or T;

a twenty-first SNP locus, wherein the twenty-first SNP locus is located at 14339957 th chromosome 4 of a cucumber reference genome, and the nucleotide base of the locus is A or G;

a twenty-second SNP locus, wherein the twenty-second SNP locus is located at 16172836 th chromosome 4 of a cucumber reference genome, and the nucleotide base of the locus is A or G;

a twenty-third SNP locus, wherein the twenty-third SNP locus is located at 20428654 th chromosome 4 of a cucumber reference genome, and the nucleotide base of the locus is G or A;

a twenty-fourth SNP locus, wherein the twenty-fourth SNP locus is located at 22774845 th chromosome 4 of a cucumber reference genome, and the nucleotide base of the locus is G or A;

a twenty-fifth SNP locus, wherein the twenty-fifth SNP locus is located at 1045863 th chromosome 5 of a cucumber reference genome, and the nucleotide base of the locus is T or G;

a twenty-sixth SNP locus, wherein the twenty-sixth SNP locus is located at 4850607 th chromosome 5 of a cucumber reference genome, and the nucleotide base of the locus is G or A;

a twenty-seventh SNP locus, wherein the twenty-seventh SNP locus is located at 5787330 th chromosome 5 of a cucumber reference genome, and the nucleotide base of the locus is T or A;

a twenty-eighth SNP locus, wherein the twenty-eighth SNP locus is located at 11869197 th chromosome 5 of a cucumber reference genome, and the nucleotide base of the locus is G or C;

a twenty-ninth SNP locus, wherein the twenty-ninth SNP locus is located at 15556517 th chromosome 5 of a cucumber reference genome, and the nucleotide base of the locus is T or G;

a thirtieth SNP locus, wherein the thirtieth SNP locus is located at 23713010 th chromosome 5 of a cucumber reference genome, and the nucleotide base of the locus is T or C;

a thirty-first SNP locus, wherein the thirty-first SNP locus is located at 24681408 th chromosome 5 of a cucumber reference genome, and the nucleotide base of the locus is C or A;

a thirty-second SNP locus, wherein the thirty-second SNP locus is located at 437035 th chromosome of the cucumber reference genome 6, and the nucleotide base of the locus is G or C;

a thirty-third SNP locus, wherein the thirty-third SNP locus is located at 6311543 th chromosome 6 of a cucumber reference genome, and the nucleotide base of the locus is T or G;

a thirty-fourth SNP locus, wherein the thirty-fourth SNP locus is located at 7644442 th chromosome 6 of a cucumber reference genome, and the nucleotide base of the locus is T or C;

a thirty-fifth SNP locus, wherein the thirty-fifth SNP locus is located at 8554160 th chromosome of a cucumber reference genome 6, and the nucleotide base of the locus is C or T;

a thirty-sixth SNP locus, wherein the thirty-sixth SNP locus is located at 9796426 th chromosome of the cucumber reference genome 6, and the nucleotide base of the locus is G or A;

a thirty-seventh SNP locus, wherein the thirty-seventh SNP locus is located at 11577012 th chromosome 6 of a cucumber reference genome, and the nucleotide base of the locus is T or G;

a thirty-eighth SNP locus, wherein the thirty-eighth SNP locus is located at 13951516 th chromosome of a cucumber reference genome 6, and the nucleotide base of the locus is T or A;

a thirty-ninth SNP locus, wherein the thirty-ninth SNP locus is located at 18948506 th chromosome 6 of a cucumber reference genome, and the nucleotide base of the locus is G or A;

a fortieth SNP locus, wherein the second SNP locus is located at 24605064 th chromosome of a 6 th chromosome of a cucumber reference genome, and the nucleotide base of the locus is A or T;

a forty-first SNP locus, wherein the fourth eleventh SNP locus is located at 4474790 th chromosome 7 of a cucumber reference genome, and the nucleotide base of the locus is C or T;

a forty-second SNP locus, wherein the fourth twelve SNP locus is located at 6777921 th chromosome 7 of a cucumber reference genome, and the nucleotide base of the locus is C or A;

a forty-third SNP locus, wherein the forty-fourth SNP locus is located at 7779724 th chromosome 7 of a cucumber reference genome, and the nucleotide base of the locus is G or A;

a forty-fourth SNP locus, wherein the forty-fourth SNP locus is located at 9945305 th chromosome 7 of a cucumber reference genome, and the nucleotide base of the locus is T or C;

a forty-fifth SNP locus, wherein the forty-fifth SNP locus is located at 10495910 th chromosome 7 of a cucumber reference genome, and the nucleotide base of the locus is A or G;

a forty-sixth SNP locus, wherein the forty-sixth SNP locus is located at 11918569 th chromosome 7 of a cucumber reference genome, and the nucleotide base of the locus is C or T;

a forty-seventh SNP locus, wherein the forty-seventh SNP locus is located at 12580918 th chromosome 7 of a cucumber reference genome, and the nucleotide base of the locus is G or A;

a forty-eighth SNP locus, wherein the forty-eighth SNP locus is located at 14166928 th chromosome 7 of a cucumber reference genome, and the nucleotide base of the locus is A or G;

the cucumber reference genome is Chinese L ong V2.

2. The SNP site according to claim 1, wherein:

the sequences of the first SNP locus and bases at the upstream and downstream are SEQ ID NO: 145 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 145 greater than or equal to 95%, 96%, 97%, 98% or 99% identity;

the sequences of the second SNP locus and bases at the upstream and downstream of the second SNP locus are SEQ ID NO: 146 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 146, greater than or equal to 95%, 96%, 97%, 98%, or 99%;

the sequences of the third SNP locus and bases at the upstream and downstream are SEQ ID NO: 147 or an idioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 147 is greater than or equal to 95%, 96%, 97%, 98% or 99% identical;

the fourth SNP locus and the sequences of the upstream and downstream bases thereof are SEQ ID NO: 148 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 148 greater than or equal to 95%, 96%, 97%, 98%, or 99%;

the fifth SNP locus and the sequences of the upstream and downstream bases thereof are SEQ ID NO: 149 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 149 is greater than or equal to 95%, 96%, 97%, 98% or 99%;

the sequences of the sixth SNP locus and the upstream and downstream bases thereof are SEQ ID NO: 150 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 150 is greater than or equal to 95%, 96%, 97%, 98%, or 99%;

the seventh SNP locus and the sequences of bases on the seventh SNP locus and upstream and downstream thereof are SEQ ID NO: 151 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 151 is greater than or equal to 95%, 96%, 97%, 98%, or 99% identical;

the sequences of the eighth SNP locus and bases at the upstream and downstream are SEQ ID NO: 152 or an interidioplasmically homologous genomic fragment thereof, more preferably to SEQ ID NO: 152 is greater than or equal to 95%, 96%, 97%, 98%, or 99% identical;

the ninth SNP site and the sequences of the upstream and downstream bases thereof are SEQ ID NO: 153 or an idioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 153 by greater than or equal to 95%, 96%, 97%, 98% or 99%;

the tenth SNP site and the sequences of the upstream and downstream bases thereof are SEQ ID NO: 154 or an interidioplasmically homologous genomic fragment thereof, more preferably to SEQ ID NO: 154 greater than or equal to 95%, 96%, 97%, 98%, or 99%;

the sequence of the eleventh SNP site and bases on the eleventh SNP site is SEQ ID NO: 155 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 155 greater than or equal to 95%, 96%, 97%, 98%, or 99%;

the sequence of the twelfth SNP site and the upstream and downstream bases thereof is SEQ ID NO: 156 or an idioplasmically homologous genomic fragment thereof, more preferably to SEQ ID NO: 156 is greater than or equal to 95%, 96%, 97%, 98%, or 99%;

the thirteenth SNP site and the sequences of the upstream and downstream bases thereof are SEQ ID NO: 157 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 157 greater than or equal to 95%, 96%, 97%, 98%, or 99%;

the sequence of the fourteenth SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 158 or an interidioplasmically homologous genomic fragment thereof, more preferably to SEQ ID NO: 158 is greater than or equal to 95%, 96%, 97%, 98% or 99% identical;

the sequence of the fifteenth SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 159 or an interidioplasmically homologous genomic fragment thereof, more preferably a fragment of SEQ ID NO: 159 is greater than or equal to 95%, 96%, 97%, 98% or 99%;

the sequence of the sixteenth SNP locus and bases on the sixteenth SNP locus is SEQ ID NO: 160 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 160, greater than or equal to 95%, 96%, 97%, 98%, or 99%;

the sequence of the seventeenth SNP site and the upstream and downstream bases thereof is SEQ ID NO: 161 or an interidioplasmic homologous genomic fragment thereof, more preferably identical to SEQ ID NO: 161 is greater than or equal to 95%, 96%, 97%, 98% or 99% identical;

the sequence of the eighteenth SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 162 or an idioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 162, greater than or equal to 95%, 96%, 97%, 98%, or 99%;

the nineteenth SNP site and the sequences of bases on the nineteenth SNP site are SEQ ID NO: 163 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 163, greater than or equal to 95%, 96%, 97%, 98%, or 99%;

the twenty-second SNP site and the sequences of bases on the twenty-second SNP site are SEQ ID NO: 164 or an interidioplasmically homologous genomic fragment thereof, more preferably to the sequence set forth in SEQ ID NO: 164 is greater than or equal to 95%, 96%, 97%, 98% or 99% identical;

the twenty-first SNP locus and the sequences of bases on the twenty-first SNP locus and bases on the twenty-first SNP locus are SEQ ID NO: 165 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 165 greater than or equal to 95%, 96%, 97%, 98% or 99% identity;

the sequence of the second twelve SNP locus and the base sequences of the second twelve SNP locus are SEQ ID NO: 166 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 166 is greater than or equal to 95%, 96%, 97%, 98%, or 99% identical;

the sequence of the twenty-third SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 167 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 167 nucleotide sequence identity of greater than or equal to 95%, 96%, 97%, 98%, or 99%;

the sequence of the twenty-fourth SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 168 or an interidioplasmic homologous genomic fragment thereof, more preferably to the sequence set forth in SEQ ID NO: 168, greater than or equal to 95%, 96%, 97%, 98%, or 99%;

the twenty-fifth SNP locus and the sequences of bases on the twenty-fifth SNP locus and the upstream and downstream of the twenty-fifth SNP locus are SEQ ID NO: 169 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 169 greater than or equal to 95%, 96%, 97%, 98%, or 99% identity in the nucleotide sequence;

the twenty-sixth SNP locus and the sequences of bases on the twenty-sixth SNP locus and bases on the twenty-sixth SNP locus are SEQ ID NO: 170 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 170, greater than or equal to 95%, 96%, 97%, 98%, or 99%;

the twenty-seventh SNP locus and the sequences of bases on the twenty-seventh SNP locus and the upstream and downstream of the twenty-seventh SNP locus are SEQ ID NO: 171 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 171 is greater than or equal to 95%, 96%, 97%, 98%, or 99% identical;

the sequences of the twenty-eight SNP locus and the upstream and downstream bases thereof are SEQ ID NO: 172 or an interidioplasmically homologous genomic fragment thereof, more preferably identical to SEQ ID NO: 172 is greater than or equal to 95%, 96%, 97%, 98%, or 99% identical;

the twenty-ninth SNP locus and the sequences of bases on the twenty-ninth SNP locus are SEQ ID NO: 173 or an interidioplasmically homologous genomic fragment thereof, more preferably to SEQ ID NO: 173, greater than or equal to 95%, 96%, 97%, 98%, or 99%;

the thirty-third SNP site and the sequences of the upstream and downstream bases thereof are SEQ ID NO: 174 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 174, or greater than or equal to 95%, 96%, 97%, 98%, or 99%;

the sequence of the thirty-first SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 175 or an idioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 175 greater than or equal to 95%, 96%, 97%, 98%, or 99%;

the sequences of the third twelve SNP loci and bases at the upper and lower ends of the third twelve SNP loci are SEQ ID NO: 176 or an interidioplasmic homologous genomic fragment thereof, more preferably identical to SEQ ID NO: 176, greater than or equal to 95%, 96%, 97%, 98%, or 99%;

the sequence of the thirteenth SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 177 or an idioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 177 greater than or equal to 95%, 96%, 97%, 98%, or 99% identity;

the sequence of the thirty-fourth SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 178 or an idioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 178 greater than or equal to 95%, 96%, 97%, 98%, or 99%;

the sequence of the fifteenth SNP locus and the bases at the upstream and downstream of the fifteenth SNP locus is SEQ ID NO: 179 or an interidioplasmically homologous genomic fragment thereof, more preferably to SEQ ID NO: 179 greater than or equal to 95%, 96%, 97%, 98% or 99% identity;

the sequences of the sixteenth SNP locus and bases on the sixteenth SNP locus are SEQ ID NO: 180 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 180 greater than or equal to 95%, 96%, 97%, 98% or 99%;

the sequences of the seventeenth SNP site and the upstream and downstream bases thereof are SEQ ID NO: 181 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 181 greater than or equal to 95%, 96%, 97%, 98%, or 99% identity;

the sequences of the third eighteen SNP locus and the upstream and downstream bases thereof are SEQ ID NO: 182 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 182 is greater than or equal to 95%, 96%, 97%, 98%, or 99% identical;

the sequence of the thirty-ninth SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 183 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 183 greater than or equal to 95%, 96%, 97%, 98% or 99% identity;

the sequence of the fortieth SNP locus and bases on the fortieth SNP locus are SEQ ID NO: 184 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 184 is greater than or equal to 95%, 96%, 97%, 98% or 99% identical;

the sequence of the fourth eleventh SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 185 or an idioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 185 is greater than or equal to 95%, 96%, 97%, 98% or 99%;

the sequence of the fourth twelve SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 186 or an idioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 186 nucleotide sequence identity of greater than or equal to 95%, 96%, 97%, 98%, or 99%;

the sequence of the fourth and the thirteenth SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 187 or an interidioplasmic homologous genomic fragment thereof, more preferably to the sequence set forth in SEQ ID NO: 187 greater than or equal to 95%, 96%, 97%, 98% or 99% identity;

the sequence of the fourteenth SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 188 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 188 is greater than or equal to 95%, 96%, 97%, 98%, or 99%;

the sequence of the fifteenth SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 189 or an intergenic homologous genomic fragment thereof, more preferably to SEQ ID NO: 189 is greater than or equal to 95%, 96%, 97%, 98% or 99% identical;

the sequence of the sixteenth SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 190 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 190 is greater than or equal to 95%, 96%, 97%, 98% or 99%;

the sequence of the fourth seventeen SNP site and the upstream and downstream bases thereof is SEQ ID NO: 191 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 191, greater than or equal to 95%, 96%, 97%, 98% or 99%;

the sequence of the fourth eighteenth SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 192 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 192 of greater than or equal to 95%, 96%, 97%, 98% or 99%.

3. Core SNP primer set for identifying cucumber germplasm authenticity, for amplifying the SNP sites of claim 1, respectively, comprising:

a first SNP primer set for amplifying the first SNP site; a second SNP primer set for amplifying the second SNP site; a third SNP primer set for amplifying the third SNP site; a fourth SNP primer set for amplifying the fourth SNP site; a fifth SNP primer set for amplifying the fifth SNP site; a sixth SNP primer set for amplifying the sixth SNP site; a seventh SNP primer set for amplifying the seventh SNP site; an eighth SNP primer set for amplifying the eighth SNP site; a ninth SNP primer set for amplifying the ninth SNP site; a tenth SNP primer set for amplifying the tenth SNP site; an eleventh SNP primer set for amplifying the eleventh SNP site; a twelfth SNP primer set for amplifying the twelfth SNP site; a thirteenth SNP primer set for amplifying the thirteenth SNP site; a fourteenth SNP primer set for amplifying the fourteenth SNP site; a fifteenth SNP primer set for amplifying the fifteenth SNP site; a sixteenth SNP primer set for amplifying the sixteenth SNP site; a seventeenth SNP primer set for amplifying the seventeenth SNP site; an eighteenth SNP primer set for amplifying the eighteenth SNP site; a nineteenth SNP primer set for amplifying the nineteenth SNP site; a twentieth SNP primer set for amplifying the twentieth SNP site; a twenty-first SNP primer set for amplifying the twenty-first SNP site; a second twelve SNP primer set for amplifying the second twelve SNP sites; a twenty-third SNP primer set for amplifying the twenty-third SNP site; a twenty-fourth SNP primer set for amplifying the twenty-fourth SNP site; a twenty-fifth SNP primer set for amplifying the twenty-fifth SNP site; a twenty-sixth SNP primer set for amplifying the twenty-sixth SNP site; a twenty-seventh SNP primer set for amplifying the twenty-seventh SNP site; a second eighteen SNP primer set for amplifying the second eighteen SNP site; a twenty-ninth SNP primer set for amplifying the twenty-ninth SNP site; a thirtieth SNP primer set for amplifying the thirtieth SNP site; a thirty-first SNP primer set for amplifying the thirty-first SNP site; a third twelve SNP primer set for amplifying the third twelve SNP sites; a thirteenth SNP primer set for amplifying the thirteenth SNP site; a thirty-fourth SNP primer set for amplifying the thirty-fourth SNP site; a fifteenth SNP primer set for amplifying the fifteenth SNP site; a sixteenth SNP primer set for amplifying the sixteenth SNP site; a third seventeen SNP primer set for amplifying the third seventeen SNP site; a third eighteen SNP primer set for amplifying the third eighteen SNP site; a thirty-ninth SNP primer set for amplifying the thirty-ninth SNP site; a fortieth SNP primer set for amplifying the fortieth SNP site; a fourth eleventh SNP primer set for amplifying the fourth eleventh SNP site; a fourth twelve SNP primer set for amplifying the fourth twelve SNP sites; a thirteenth SNP primer set for amplifying the thirteenth SNP site; a fourteenth SNP primer set for amplifying the fourteenth SNP site; a fifteenth SNP primer set for amplifying the fifteenth SNP site; a sixteenth SNP primer set for amplifying the sixteenth SNP site; a seventeenth SNP primer set for amplifying the seventeenth SNP site; a forty-eighth SNP primer set for amplifying the forty-eighth SNP site.

4. The SNP primer set according to claim 3, wherein:

the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer of the first SNP primer set are respectively matched with the sequence shown in SEQ ID NO: 1. SEQ ID NO: 2. SEQ ID NO: 3 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

and the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer of the second SNP primer set are respectively matched with the sequence shown in SEQ ID NO: 4. SEQ ID NO: 5. SEQ ID NO: 6 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

and the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer of the third SNP primer set are respectively matched with the sequences shown in SEQ ID NO: 7. SEQ ID NO: 8. SEQ ID NO: 9 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

and the fourth SNP primer group, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequences shown in SEQ ID NO: 10. SEQ ID NO: 11. SEQ ID NO: 12 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

and in the fifth SNP primer group, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequences shown in SEQ ID NO: 13. SEQ ID NO: 14. SEQ ID NO: 15, is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98%, or 99%, preferably 100%;

and in the sixth SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequences shown in SEQ ID NO: 16. SEQ ID NO: 17. SEQ ID NO: 18, is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98%, or 99%, preferably 100%;

and in the seventh SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequences shown in SEQ ID NO: 19. SEQ ID NO: 20. SEQ ID NO: 21 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

and in the eighth SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequences shown in SEQ ID NO: 22. SEQ ID NO: 23. SEQ ID NO: 24 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

and in the ninth SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequences shown in SEQ ID NO: 25. SEQ ID NO: 26. SEQ ID NO: 27 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

and in the tenth SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequence shown in SEQ ID NO: 28. SEQ ID NO: 29. SEQ ID NO: 30 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

and the eleventh SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequence shown in SEQ ID NO: 31. SEQ ID NO: 32. SEQ ID NO: 33, is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98%, or 99%, preferably 100%;

and the twelfth SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequence shown in SEQ ID NO: 34. SEQ ID NO: 35. SEQ ID NO: 36 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

and in the thirteenth SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with SEQ ID NO: 37. SEQ ID NO: 38. SEQ ID NO: 39, greater than or equal to 85%, 90%, 95%, 96%, 97%, 98%, or 99%, preferably 100%;

and in the fourteenth SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequence shown in SEQ ID NO: 40. SEQ ID NO: 41. SEQ ID NO: 42 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

and in the fifteenth SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequence shown in SEQ ID NO: 43. SEQ ID NO: 44. SEQ ID NO: 45 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

and in the sixteenth SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively connected with SEQ ID NO: 46. SEQ ID NO: 47. SEQ ID NO: 48, is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98%, or 99%, preferably 100%;

and in the seventeenth SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequences shown in SEQ ID NO: 49. SEQ ID NO: 50. SEQ ID NO: 51, is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98%, or 99%, preferably 100%;

the eighteenth SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively connected with SEQ ID NO: 52. SEQ ID NO: 53. SEQ ID NO: 54 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

and in the nineteenth SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequences shown in SEQ ID NO: 55. SEQ ID NO: 56. SEQ ID NO: 57 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

and the twentieth SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequence shown in SEQ ID NO: 58. SEQ ID NO: 59. SEQ ID NO: 60, preferably 100%, or greater than 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity;

the twenty-first SNP primer group, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively connected with the sequence shown in SEQ ID NO: 61. SEQ ID NO: 62. SEQ ID NO: 63 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

and the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer of the second twelve SNP primer set are respectively matched with the sequence shown in SEQ ID NO: 64. SEQ ID NO: 65. SEQ ID NO: 66 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

and in the twenty-third SNP primer group, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequence shown in SEQ ID NO: 67. SEQ ID NO: 68. SEQ ID NO: 69 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

and in the twenty-fourth SNP primer group, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequence shown in SEQ ID NO: 70. SEQ ID NO: 71. SEQ ID NO: 72 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

and in the twenty-fifth SNP primer group, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively connected with the primers shown in SEQ ID NO: 73. SEQ ID NO: 74. SEQ ID NO: 75 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

and in the twenty-sixth SNP primer group, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively connected with the primers shown in SEQ ID NO: 76. SEQ ID NO: 77. SEQ ID NO: 78, is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98%, or 99%, preferably 100%;

and in the twenty-seventh SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively connected with SEQ ID NO: 79. SEQ ID NO: 80. SEQ ID NO: 81 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

and the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer of the second eighteen SNP primer set are respectively connected with the sequences shown in SEQ ID NO: 82. SEQ ID NO: 83. SEQ ID NO: 84 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

and in the twenty-ninth SNP primer group, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively connected with SEQ ID NO: 85. SEQ ID NO: 86. SEQ ID NO: 87 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

and in the thirtieth SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequence shown in SEQ ID NO: 88. SEQ ID NO: 89. SEQ ID NO: 90 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

and in the thirty-first SNP primer group, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequence shown in SEQ ID NO: 91. SEQ ID NO: 92. SEQ ID NO: 93 greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

and in the third twelve SNP primer group, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequences shown in SEQ ID NO: 94. SEQ ID NO: 95. SEQ ID NO: 96 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

and in the thirteenth SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequence shown in SEQ ID NO: 97. SEQ ID NO: 98. SEQ ID NO: 99, and preferably 100%, or greater than 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity;

and in the thirty-fourth SNP primer group, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequence shown in SEQ ID NO: 100. SEQ ID NO: 101. SEQ ID NO: 102 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

and in the thirty-fifth SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequences shown in SEQ ID NO: 103. SEQ ID NO: 104. SEQ ID NO: 105 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

and in the sixteenth SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively connected with the primers shown in SEQ ID NO: 106. SEQ ID NO: 107. SEQ ID NO: 108 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

and in the third seventeen SNP primer group, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequences shown in SEQ ID NO: 109. SEQ ID NO: 110. SEQ ID NO: 111 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

and in the third eighteen SNP primer group, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively connected with the sequences shown in SEQ ID NO: 112. SEQ ID NO: 113. SEQ ID NO: 114 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

and in the third nineteenth SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequences shown in SEQ ID NO: 115. SEQ ID NO: 116. SEQ ID NO: 117 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

and the fortieth SNP primer group, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequence shown in SEQ ID NO: 118. SEQ ID NO: 119. SEQ ID NO: 120, is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98%, or 99%, preferably 100%;

and in the fourth eleventh SNP primer group, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequence shown in SEQ ID NO: 121. SEQ ID NO: 122. SEQ ID NO: 123 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

and in the fourth twelve SNP primer group, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequences shown in SEQ ID NO: 124. SEQ ID NO: 125. SEQ ID NO: 126, is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98%, or 99%, preferably 100%;

and in the fourth and the thirteenth SNP primer sets, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequences shown in SEQ ID NO: 127. SEQ ID NO: 128. SEQ ID NO: 129 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

and in the fourteenth SNP primer group, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequence shown in SEQ ID NO: 130. SEQ ID NO: 131. SEQ ID NO: 132 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

and in the fifteenth SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequence shown in SEQ ID NO: 133. SEQ ID NO: 134. SEQ ID NO: 135 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

and in the sixteenth SNP primer group, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively connected with the primers shown in SEQ ID NO: 136. SEQ ID NO: 137. SEQ ID NO: 138 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

and in the fourth seventeen SNP primer group, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequences shown in SEQ ID NO: 139. SEQ ID NO: 140. SEQ ID NO: 141, greater than or equal to 85%, 90%, 95%, 96%, 97%, 98%, or 99%, preferably 100%;

and in the fourth eighteenth SNP primer group, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively connected with the primers shown in SEQ ID NO: 142. SEQ ID NO: 143. SEQ ID NO: 144 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

preferably, the first and second upstream primers in each set of primers are linked to different fluorescent molecules, more preferably, the fluorescent molecules are selected from ROX, TAMRA, FAM, HEX.

5. The core SNP kit for identifying the authenticity of cucumber germplasm is characterized in that: the SNP kit is prepared into a competitive allele specificity PCR reaction system; the reaction system comprises:

the SNP primer set according to claim 3 or 4,

preferably, in the SNP primer sets, the concentration ratio of the first upstream primer, the second upstream primer and the downstream primer of each primer set in the system is 2:2: 5.

6. A cucumber germplasm DNA fingerprint database based on core SNP markers is characterized in that: the DNA fingerprint database includes: genotype of a standard cucumber germplasm at a SNP site according to claim 1.

7. The DNA fingerprint database of claim 6, wherein: the standard cucumber germplasm is selected from the following 105 cucumber germplasms:

beijing lesser spiny melon, CM8537, Focus pinus, Green Tangerine Peel octabifurcate, Roqing 7314-2-6-1-1, Liaotong dense thorn, yellow iron lesser spiny melon, bvrc, Cucumis sativus, leaf tribe, Liaoyang leaf tribe, Xintai dense thorn, bvrc, bvrc, bvrc, Qingdao autumn leaf tribe, KagaFushinari, Sagami Hanpakuri, Fushinari Kyuri, anonga Suyo Kyuri, uda, Sakatatatsusago, Honshu Aibai, Sekino No.2(Ochiai No.2), CGN19828, bvrc, Cucumis sativus, autumn cucumber, dew, bvlon, bvrc, bvrc, bvrc, Sichuan white melon, Bvre, Btyrc, Purch No.2, Kanrc, Purch No.2, Karcuit No.2, Kvkuri, Kyu Ka Karvi, Karvi Kyu Kanvex, Kanvicu Kyu, Kanvicu, Karcu, Karcuit, Karcu, Karve, Ka, Karve.

8. The method of constructing a DNA fingerprint database according to claim 6, wherein: the construction method comprises the following steps:

and (3) PCR reaction steps: carrying out competitive allele specific PCR amplification reaction on standard cucumber germplasm by adopting the PCR reaction system as claimed in claim 5 to obtain a PCR reaction product;

SNP locus genotype obtaining step: detecting the PCR reaction product to obtain the genotype of the SNP locus;

preferably, the detection is fluorescence signal detection or direct sequencing.

9. A detection method for identifying the authenticity of cucumber germplasm is characterized by comprising the following steps: the detection method comprises the following steps:

the method comprises the following steps: detecting the genotype of the SNP locus of cucumber to be detected according to claim 1;

step two: and (3) germplasm judgment of the cucumber to be detected:

if the number of the genotype of the cucumber to be detected based on the 48 SNP loci and the number of the different loci of a specified germplasm in the standard cucumber germplasm in the database of claim 6 or 7 based on the genotype of the 48 SNP loci are 0-2, the cucumber to be detected is judged as a similar germplasm with the specified germplasm;

if the number of the genotype of the cucumber to be detected based on the 48 SNP loci and the number of the different loci of a specified germplasm in the standard cucumber germplasm based on the genotype of the 48 SNP loci in the database of claim 6 or 7 are more than 2, the cucumber to be detected and the specified germplasm are judged to be different cucumber germplasms;

preferably, the result of the determination is obtained from a cluster analysis.

10. The SNP site according to claim 1 or 2, or the SNP primer combination according to claim 3 or 4, or the SNP kit according to claim 5, or the DNA fingerprint database according to claim 6 or 7, or the DNA fingerprint database obtained by the construction method according to claim 8, or the detection method according to claim 9, wherein the SNP site is used in the following X1 or X2:

x1: identifying whether the germplasm of the cucumber to be detected belongs to one of standard cucumber germplasms;

x2: and identifying the specific germplasm of the cucumber to be detected as the standard cucumber germplasm.

Technical Field

The invention belongs to the field of molecular markers and detection thereof, and particularly relates to a core SNP (single nucleotide polymorphism) site and primer combination for identifying the authenticity of cucumber germplasm, a DNA (deoxyribonucleic acid) fingerprint database and database construction method based on the SNP site, and an identification method and application of the authenticity of cucumber germplasm based on the SNP site.

Background

Germplasm resources are the foundation of breeding. The nineteenth article of the seed method states that germplasm resources refer to a basic material for breeding new varieties of plants, including propagation materials of cultivars and wild species of various plants, and genetic materials of various plants artificially created using the propagation materials. The germplasm resource is an important natural resource formed by natural evolution and artificial creation, accumulates extremely rich genetic variation, and is a material basis for breeding new varieties and developing agricultural production by human beings. The discovery and utilization of germplasm resources determine the key to the breakthrough development of modern crop breeding. Therefore, the identification and protection of germplasm resources are more important in the aspects of identifying authenticity and optimizing sources compared with specific varieties. However, at present, the identification and protection work of most germplasm resources is in a blank state.

The cucumber is one of the vegetable crops which are popular with Chinese consumers due to the unique flavor of the cucumber. In 2018, FAOSTAT promulgates its yield in China of 5629.4 tons (http:// www.fao.org/FAOSTAT/zh/# data/QC), accounting for 70% of the total cucumber yield in the world. At present, the cucumber varieties in the market are rich, but most of the cucumber varieties are hybrid varieties. The registered varieties of cucumber published by the ministry of agriculture are thousands of, and the hybrid varieties are more than 95%. The cucumber germplasm resource is genetic material for cultivating cucumber hybrid, the genetic characteristics of the cucumber germplasm resource directly determine the yield and quality of the hybrid, however, research on cucumber germplasm resource protection is not reported. As is known, a cucumber germplasm with excellent characters needs to be cultivated by a breeder for years, but illegal molecules are stolen, so that the germplasm intellectual property of the breeder is seriously infringed, the enthusiasm of the breeder is influenced, and the benign development of the cucumber breeding industry is hindered. At present, a molecular marker detection method is applied to identification of cucumber varieties, but researches on identification and protection of cucumber germplasm resources (breeding parents) are rarely involved, so that the phenomenon of stealing cucumber germplasm is serious, a plurality of similar germplasm are generated, and the protection of intellectual property rights of the cucumber germplasm is hindered. Meanwhile, the cucumber variety cultivated by using the similar cucumber germplasm also brings certain difficulty to the registration and the approval of the cucumber variety, and influences the supervision of the cucumber variety. Therefore, a set of DNA rapid detection technology suitable for identifying the cucumber germplasm is urgently needed to be developed.

Cucumber germplasm is divided into 4 major classes, including the indian, sidsna, european and eastern subtypes. Wherein, the Indian type is cucumber wild germplasm; the Xishuangbanna type is the main type of cucumber planted in the Xishuangbanna area in China; european and eastern subtypes are the main germplasm for cultivating cucumber varieties in China. At present, more than three thousand parts of cucumber germplasm in the world are collected by national research institutions, but the research has not reported that the cucumber germplasm is identified by using SNP loci, so that a cucumber DNA fingerprint database based on core SNP loci needs to be constructed, and a method and application for identifying the cucumber germplasm based on the core SNP loci are also needed.

Disclosure of Invention

The invention provides a core SNP locus and a primer combination for identifying cucumber germplasm, a DNA fingerprint database and database construction method based on the SNP locus, and an identification method and application of the cucumber germplasm based on the SNP locus.

The invention is realized by the following technical scheme:

the genomic DNA of cucumber is divided into a fifteenth genomic DNA, a fourteenth genomic DNA, a fifteenth genomic DNA, a fifteenth genomic DNA, a fifteenth genomic DNA, a genomic DNA, a genomic DNA, a genome, a genome, a genome, a genome, a genome, a.

In some embodiments, the sequence of the first SNP site and its upstream and downstream bases is SEQ ID NO: 145 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 145 greater than or equal to 95%, 96%, 97%, 98% or 99% identity; the sequences of the second SNP locus and bases at the upstream and downstream of the second SNP locus are SEQ ID NO: 146 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 146, greater than or equal to 95%, 96%, 97%, 98%, or 99%; the sequences of the third SNP locus and bases at the upstream and downstream are SEQ ID NO: 147 or an interidioplasmic homologous genomic fragment thereof, more preferably a fragment identical to SEQ id no: 147 is greater than or equal to 95%, 96%, 97%, 98% or 99% identical; the fourth SNP locus and the sequences of the upstream and downstream bases thereof are SEQ ID NO: 148 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 148 greater than or equal to 95%, 96%, 97%, 98%, or 99%; the fifth SNP locus and the sequences of the upstream and downstream bases thereof are SEQ ID NO: 149 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 149 is greater than or equal to 95%, 96%, 97%, 98% or 99%; the sequences of the sixth SNP locus and the upstream and downstream bases thereof are SEQ ID NO: 150 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 150 is greater than or equal to 95%, 96%, 97%, 98%, or 99%; the seventh SNP locus and the sequences of bases on the seventh SNP locus and upstream and downstream thereof are SEQ ID NO: 151 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 151 is greater than or equal to 95%, 96%, 97%, 98%, or 99% identical; the sequences of the eighth SNP locus and bases at the upstream and downstream are SEQ ID NO: 152 or an interidioplasmically homologous genomic fragment thereof, more preferably to SEQ ID NO: 152 is greater than or equal to 95%, 96%, 97%, 98%, or 99% identical; the ninth SNP site and the sequences of the upstream and downstream bases thereof are SEQ ID NO: 153 or an idioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 153 by greater than or equal to 95%, 96%, 97%, 98% or 99%; the tenth SNP site and the sequences of the upstream and downstream bases thereof are SEQ ID NO: 154 or an interidioplasmically homologous genomic fragment thereof, more preferably to SEQ ID NO: 154 greater than or equal to 95%, 96%, 97%, 98%, or 99%; the sequence of the eleventh SNP site and bases on the eleventh SNP site is SEQ ID NO: 155 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 155 greater than or equal to 95%, 96%, 97%, 98%, or 99%; the sequence of the twelfth SNP site and the upstream and downstream bases thereof is SEQ ID NO: 156 or an idioplasmically homologous genomic fragment thereof, more preferably to SEQ ID NO: 156 is greater than or equal to 95%, 96%, 97%, 98%, or 99%; the thirteenth SNP site and the sequences of the upstream and downstream bases thereof are SEQ ID NO: 157 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 157 greater than or equal to 95%, 96%, 97%, 98%, or 99%; the sequence of the fourteenth SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 158 or an interidioplasmically homologous genomic fragment thereof, more preferably to SEQ ID NO: 158 is greater than or equal to 95%, 96%, 97%, 98% or 99% identical; the sequence of the fifteenth SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 159 or an interidioplasmically homologous genomic fragment thereof, more preferably a fragment of SEQ ID NO: 159 is greater than or equal to 95%, 96%, 97%, 98% or 99%; the sequence of the sixteenth SNP locus and bases on the sixteenth SNP locus is SEQ ID NO: 160 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 160, greater than or equal to 95%, 96%, 97%, 98%, or 99%; the sequence of the seventeenth SNP site and the upstream and downstream bases thereof is SEQ ID NO: 161 or an interidioplasmic homologous genomic fragment thereof, more preferably to seq id NO: 161 is greater than or equal to 95%, 96%, 97%, 98% or 99% identical; the sequence of the eighteenth SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 162 or an idioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 162, greater than or equal to 95%, 96%, 97%, 98%, or 99%; the nineteenth SNP site and the sequences of bases on the nineteenth SNP site are SEQ ID NO: 163 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 163, greater than or equal to 95%, 96%, 97%, 98%, or 99%; the twenty-second SNP site and the sequences of bases on the twenty-second SNP site are SEQ ID NO: 164 or an interidioplasmically homologous genomic fragment thereof, more preferably to the sequence set forth in SEQ ID NO: 164 is greater than or equal to 95%, 96%, 97%, 98% or 99% identical; the twenty-first SNP locus and the sequences of bases on the twenty-first SNP locus and bases on the twenty-first SNP locus are SEQ ID NO: 165 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 165 greater than or equal to 95%, 96%, 97%, 98% or 99% identity; the sequence of the second twelve SNP locus and the base sequences of the second twelve SNP locus are SEQ ID NO: 166 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 166 is greater than or equal to 95%, 96%, 97%, 98%, or 99% identical; the sequence of the twenty-third SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 167 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 167 nucleotide sequence identity of greater than or equal to 95%, 96%, 97%, 98%, or 99%; the sequence of the twenty-fourth SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 168 or an interidioplasmic homologous genomic fragment thereof, more preferably to the sequence set forth in SEQ ID NO: 168, greater than or equal to 95%, 96%, 97%, 98%, or 99%; the twenty-fifth SNP locus and the sequences of bases on the twenty-fifth SNP locus and the upstream and downstream of the twenty-fifth SNP locus are SEQ ID NO: 169 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 169 greater than or equal to 95%, 96%, 97%, 98%, or 99% identity in the nucleotide sequence; the twenty-sixth SNP locus and the sequences of bases on the twenty-sixth SNP locus and bases on the twenty-sixth SNP locus are SEQ ID NO: 170 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 170, greater than or equal to 95%, 96%, 97%, 98%, or 99%; the twenty-seventh SNP locus and the sequences of bases on the twenty-seventh SNP locus and the upstream and downstream of the twenty-seventh SNP locus are SEQ ID NO: 171 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 171 is greater than or equal to 95%, 96%, 97%, 98%, or 99% identical; the sequences of the twenty-eight SNP locus and the upstream and downstream bases thereof are SEQ ID NO: 172 or an interidioplasmic homologous genomic fragment thereof, more preferably to seq id NO: 172 is greater than or equal to 95%, 96%, 97%, 98%, or 99% identical; the twenty-ninth SNP locus and the sequences of bases on the twenty-ninth SNP locus are SEQ ID NO: 173 or an interidioplasmically homologous genomic fragment thereof, more preferably to SEQ ID NO: 173, greater than or equal to 95%, 96%, 97%, 98%, or 99%; the thirty-third SNP site and the sequences of the upstream and downstream bases thereof are SEQ ID NO: 174 or an interidioplasmic homologous genomic fragment thereof, more preferably identical to SEQ id no: 174, or greater than or equal to 95%, 96%, 97%, 98%, or 99%; the sequence of the thirty-first SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 175 or an idioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 175 greater than or equal to 95%, 96%, 97%, 98%, or 99%; the sequences of the third twelve SNP loci and bases on the third twelve SNP loci are SEQ ID NO: 176 or an interidioplasmic homologous genomic fragment thereof, more preferably identical to SEQ ID NO: 176, greater than or equal to 95%, 96%, 97%, 98%, or 99%; the sequence of the thirteenth SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 177 or an idioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 177 greater than or equal to 95%, 96%, 97%, 98%, or 99% identity; the sequence of the thirty-fourth SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 178 or an idioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 178 greater than or equal to 95%, 96%, 97%, 98%, or 99%; the sequence of the fifteenth SNP locus and the bases at the upstream and downstream of the fifteenth SNP locus is SEQ ID NO: 179 or an interidioplasmically homologous genomic fragment thereof, more preferably to SEQ ID NO: 179 greater than or equal to 95%, 96%, 97%, 98% or 99% identity; the sequences of the sixteenth SNP locus and bases on the sixteenth SNP locus are SEQ ID NO: 180 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 180 greater than or equal to 95%, 96%, 97%, 98% or 99%; the sequences of the seventeenth SNP site and the upstream and downstream bases thereof are SEQ ID NO: 181 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 181 greater than or equal to 95%, 96%, 97%, 98%, or 99% identity; the sequences of the third eighteen SNP locus and the upstream and downstream bases thereof are SEQ ID NO: 182 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 182 is greater than or equal to 95%, 96%, 97%, 98%, or 99% identical; the sequence of the thirty-ninth SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 183 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 183 greater than or equal to 95%, 96%, 97%, 98% or 99% identity; the sequence of the fortieth SNP locus and bases on the fortieth SNP locus are SEQ ID NO: 184 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 184 is greater than or equal to 95%, 96%, 97%, 98% or 99% identical; the sequence of the fourth eleventh SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 185 or an idioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 185 is greater than or equal to 95%, 96%, 97%, 98% or 99%; the sequence of the fourth twelve SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 186 or an idioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 186 nucleotide sequence identity of greater than or equal to 95%, 96%, 97%, 98%, or 99%; the sequence of the fourth and the thirteenth SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 187 or an interidioplasmic homologous genomic fragment thereof, more preferably identical to SEQ id no: 187 greater than or equal to 95%, 96%, 97%, 98% or 99% identity; the sequence of the fourteenth SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 188 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 188 is greater than or equal to 95%, 96%, 97%, 98%, or 99%; the sequence of the fifteenth SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 189 or an intergenic homologous genomic fragment thereof, more preferably to SEQ ID NO: 189 is greater than or equal to 95%, 96%, 97%, 98% or 99% identical; the sequence of the sixteenth SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 190 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 190 is greater than or equal to 95%, 96%, 97%, 98% or 99%; the sequence of the fourth seventeen SNP site and the upstream and downstream bases thereof is SEQ ID NO: 191 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 191, greater than or equal to 95%, 96%, 97%, 98% or 99%; the sequence of the fourth eighteenth SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 192 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 192 of greater than or equal to 95%, 96%, 97%, 98% or 99%.

The core SNP primer set for identifying the germplasm authenticity of the cucumber is used for respectively amplifying the SNP loci, and comprises the following components: a first SNP primer set for amplifying the first SNP site; a second SNP primer set for amplifying the second SNP site; a third SNP primer set for amplifying the third SNP site; a fourth SNP primer set for amplifying the fourth SNP site; a fifth SNP primer set for amplifying the fifth SNP site; a sixth SNP primer set for amplifying the sixth SNP site; a seventh SNP primer set for amplifying the seventh SNP site; an eighth SNP primer set for amplifying the eighth SNP site; a ninth SNP primer set for amplifying the ninth SNP site; a tenth SNP primer set for amplifying the tenth SNP site; an eleventh SNP primer set for amplifying the eleventh SNP site; a twelfth SNP primer set for amplifying the twelfth SNP site; a thirteenth SNP primer set for amplifying the thirteenth SNP site; a fourteenth SNP primer set for amplifying the fourteenth SNP site; a fifteenth SNP primer set for amplifying the fifteenth SNP site; a sixteenth SNP primer set for amplifying the sixteenth SNP site; a seventeenth SNP primer set for amplifying the seventeenth SNP site; an eighteenth SNP primer set for amplifying the eighteenth SNP site; a nineteenth SNP primer set for amplifying the nineteenth SNP site; a twentieth SNP primer set for amplifying the twentieth SNP site; a twenty-first SNP primer set for amplifying the twenty-first SNP site; a second twelve SNP primer set for amplifying the second twelve SNP sites; a twenty-third SNP primer set for amplifying the twenty-third SNP site; a twenty-fourth SNP primer set for amplifying the twenty-fourth SNP site; a twenty-fifth SNP primer set for amplifying the twenty-fifth SNP site; a twenty-sixth SNP primer set for amplifying the twenty-sixth SNP site; a twenty-seventh SNP primer set for amplifying the twenty-seventh SNP site; a second eighteen SNP primer set for amplifying the second eighteen SNP site; a twenty-ninth SNP primer set for amplifying the twenty-ninth SNP site; a thirtieth SNP primer set for amplifying the thirtieth SNP site; a thirty-first SNP primer set for amplifying the thirty-first SNP site; a third twelve SNP primer set for amplifying the third twelve SNP sites; a thirteenth SNP primer set for amplifying the thirteenth SNP site; a thirty-fourth SNP primer set for amplifying the thirty-fourth SNP site; a fifteenth SNP primer set for amplifying the fifteenth SNP site; a sixteenth SNP primer set for amplifying the sixteenth SNP site; a third seventeen SNP primer set for amplifying the third seventeen SNP site; a third eighteen SNP primer set for amplifying the third eighteen SNP site; a thirty-ninth SNP primer set for amplifying the thirty-ninth SNP site; a fortieth SNP primer set for amplifying the fortieth SNP site; a fourth eleventh SNP primer set for amplifying the fourth eleventh SNP site; a fourth twelve SNP primer set for amplifying the fourth twelve SNP sites; a thirteenth SNP primer set for amplifying the thirteenth SNP site; a fourteenth SNP primer set for amplifying the fourteenth SNP site; a fifteenth SNP primer set for amplifying the fifteenth SNP site; a sixteenth SNP primer set for amplifying the sixteenth SNP site; a seventeenth SNP primer set for amplifying the seventeenth SNP site; a forty-eighth SNP primer set for amplifying the forty-eighth SNP site.

In some embodiments, the first SNP primer set, the specific portion of the first forward primer, the specific portion of the second forward primer, and the downstream primer are each identical to SEQ ID NO: 1. SEQ ID NO: 2. SEQ ID NO: 3 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%; and the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer of the second SNP primer set are respectively matched with the sequence shown in SEQ ID NO: 4. SEQ ID NO: 5. SEQ ID NO: 6 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%; and the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer of the third SNP primer set are respectively matched with the sequences shown in SEQ ID NO: 7. SEQ ID NO: 8. SEQ ID NO: 9 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%; and the fourth SNP primer group, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequences shown in SEQ ID NO: 10. SEQ ID NO: 11. SEQ ID NO: 12 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%; and in the fifth SNP primer group, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequences shown in SEQ ID NO: 13. SEQ ID NO: 14. SEQ ID NO: 15, is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98%, or 99%, preferably 100%; and in the sixth SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequences shown in SEQ ID NO: 16. SEQ ID NO: 17. SEQ ID NO: 18, is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98%, or 99%, preferably 100%; and in the seventh SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequences shown in SEQ ID NO: 19. SEQ ID NO: 20. SEQ ID NO: 21 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%; and in the eighth SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequences shown in SEQ ID NO: 22. SEQ ID NO: 23. SEQ ID NO: 24 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%; and in the ninth SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequences shown in SEQ ID NO: 25. SEQ ID NO: 26. SEQ ID NO: 27 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%; and in the tenth SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequence shown in SEQ ID NO: 28. SEQ ID NO: 29. SEQ ID NO: 30 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%; and the eleventh SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequence shown in SEQ ID NO: 31. SEQ ID NO: 32. SEQ ID NO: 33, is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98%, or 99%, preferably 100%; and the twelfth SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequence shown in SEQ ID NO: 34. SEQ ID NO: 35. SEQ ID NO: 36 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%; and in the thirteenth SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with SEQ ID NO: 37. SEQ ID NO: 38. SEQ ID NO: 39, greater than or equal to 85%, 90%, 95%, 96%, 97%, 98%, or 99%, preferably 100%; and in the fourteenth SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequence shown in SEQ ID NO: 40. SEQ ID NO: 41. SEQ ID NO: 42 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%; and in the fifteenth SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequence shown in SEQ ID NO: 43. SEQ ID NO: 44. SEQ ID NO: 45 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%; and in the sixteenth SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively connected with SEQ ID NO: 46. SEQ ID NO: 47. SEQ ID NO: 48, is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98%, or 99%, preferably 100%; and in the seventeenth SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequences shown in SEQ ID NO: 49. SEQ ID NO: 50. SEQ ID NO: 51, is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98%, or 99%, preferably 100%; the eighteenth SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively connected with SEQ ID NO: 52. SEQ ID NO: 53. SEQ ID NO: 54 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%; and in the nineteenth SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequences shown in SEQ ID NO: 55. SEQ ID NO: 56. SEQ ID NO: 57 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%; and the twentieth SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequence shown in SEQ ID NO: 58. SEQ ID NO: 59. SEQ ID NO: 60, preferably 100%, or greater than 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity; the twenty-first SNP primer group, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively connected with the sequence shown in SEQ ID NO: 61. SEQ ID NO: 62. SEQ ID NO: 63 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%; and the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer of the second twelve SNP primer set are respectively matched with the sequence shown in SEQ ID NO: 64. SEQ ID NO: 65. SEQ ID NO: 66 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%; and in the twenty-third SNP primer group, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively connected with the primers shown in SEQ ID NO: 67. SEQ ID NO: 68. SEQ ID NO: 69 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%; and in the twenty-fourth SNP primer group, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequence shown in SEQ ID NO: 70. SEQ ID NO: 71. SEQ ID NO: 72 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%; and in the twenty-fifth SNP primer group, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively connected with the primers shown in SEQ ID NO: 73. SEQ ID NO: 74. SEQ ID NO: 75 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%; and in the twenty-sixth SNP primer group, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively connected with the primers shown in SEQ ID NO: 76. SEQ ID NO: 77. SEQ ID NO: 78, is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98%, or 99%, preferably 100%; and in the twenty-seventh SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively connected with SEQ ID NO: 79. SEQ ID NO: 80. SEQ ID NO: 81 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%; and the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer of the second eighteen SNP primer set are respectively connected with the sequences shown in SEQ ID NO: 82. SEQ ID NO: 83. SEQ ID NO: 84 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%; and in the twenty-ninth SNP primer group, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively connected with SEQ ID NO: 85. SEQ ID NO: 86. SEQ ID NO: 87 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%; and in the thirtieth SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequence shown in SEQ ID NO: 88. SEQ ID NO: 89. SEQ ID NO: 90 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%; and in the thirty-first SNP primer group, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequence shown in SEQ ID NO: 91. SEQ ID NO: 92. SEQ ID NO: 93 greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%; and in the third twelve SNP primer group, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively connected with the sequences shown in SEQ ID NOs in the sequence table: 94. SEQ ID NO: 95. SEQ ID NO: 96 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%; and in the thirteenth SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequence shown in SEQ ID NO: 97. SEQ ID NO: 98. SEQ ID NO: 99, and preferably 100%, or greater than 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity; and in the thirty-fourth SNP primer group, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequence shown in SEQ ID NO: 100. SEQ ID NO: 101. SEQ ID NO: 102 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%; and in the thirty-fifth SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequences shown in SEQ ID NO: 103. SEQ ID NO: 104. SEQ ID NO: 105 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%; and in the sixteenth SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively connected with the primers shown in SEQ ID NO: 106. SEQ ID NO: 107. SEQ ID NO: 108 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%; and in the third seventeen SNP primer group, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively connected with the primers shown in SEQ ID NO: 109. SEQ ID NO: 110. SEQ ID NO: 111 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%; and in the third eighteen SNP primer group, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively connected with the sequences shown in SEQ ID NO: 112. SEQ ID NO: 113. SEQ ID NO: 114 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%; and in the third nineteenth SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequences shown in SEQ ID NO: 115. SEQ ID NO: 116. SEQ ID NO: 117 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%; and the fortieth SNP primer group, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequence shown in SEQ ID NO: 118. SEQ ID NO: 119. SEQ ID NO: 120, is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98%, or 99%, preferably 100%; and in the fourth eleventh SNP primer group, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequence shown in SEQ ID NO: 121. SEQ ID NO: 122. SEQ ID NO: 123 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%; and in the fourth twelve SNP primer group, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively connected with the sequence shown in SEQ ID NO: 124. SEQ ID NO: 125. SEQ ID NO: 126, is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98%, or 99%, preferably 100%; and in the fourth and the thirteenth SNP primer sets, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequences shown in SEQ ID NO: 127. SEQ ID NO: 128. SEQ ID NO: 129 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%; and in the fourteenth SNP primer group, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequence shown in SEQ ID NO: 130. SEQ ID NO: 131. SEQ ID NO: 132 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%; and in the fifteenth SNP primer set, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively matched with the sequence shown in SEQ ID NO: 133. SEQ ID NO: 134. SEQ ID NO: 135 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%; and in the sixteenth SNP primer group, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively connected with the primers shown in SEQ ID NO: 136. SEQ ID NO: 137. SEQ ID NO: 138 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%; and in the fourth seventeen SNP primer group, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively connected with the primers shown in SEQ ID NO: 139. SEQ ID NO: 140. SEQ ID NO: 141, greater than or equal to 85%, 90%, 95%, 96%, 97%, 98%, or 99%, preferably 100%; and in the fourth eighteenth SNP primer group, the specific part of the first upstream primer, the specific part of the second upstream primer and the downstream primer are respectively connected with the primers shown in SEQ ID NO: 142. SEQ ID NO: 143. SEQ ID NO: 144 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%; preferably, the first and second upstream primers in each set of primers are linked to different fluorescent molecules, more preferably, the fluorescent molecules are selected from ROX, TAMRA, FAM, HEX.

The core SNP kit for identifying the germplasm authenticity of the cucumber is prepared into a competitive allele specificity PCR reaction system; the reaction system comprises: in the above SNP primer sets, the concentration ratio of the first upstream primer, the second upstream primer and the downstream primer of each primer set in the SNP primer set is preferably 2:2: 5.

A cucumber germplasm DNA fingerprint database based on core SNP markers, the DNA fingerprint database comprising: and (3) the genotype of the SNP locus of the standard cucumber germplasm.

In some embodiments, the standard cucumber germplasm is selected from the group consisting of 105 cucumber germplasm, including Peking spiny cucumber, CM8537, Fugu, Mitsui, Mitsu, Sauta Natsungo, Honshu Aibai, Sekino No.2 (Ochiaai No.2), CGN19828, bvsra, Daxu, autumn, Cold dew, WCbncy, Barr, Querc, Sekino No.2(Ochiai No.2), cucumber Bvru, Mitsu, Mitsui, Mitsukura, Mitsukui, Mitsukura, Mitsu kura, Mitsu kui, Mitsui, Mitsukui, Mitsui No.2(Ochiai, Mitsui, Mitsu.

The construction method of the DNA fingerprint database comprises the following steps: and (3) PCR reaction steps: carrying out competitive allele specificity PCR amplification reaction on the standard cucumber germplasm by adopting the PCR reaction system to obtain a PCR reaction product; SNP locus genotype obtaining step: detecting the PCR reaction product to obtain the genotype of the SNP locus; preferably, the detection is fluorescence signal detection or direct sequencing.

A detection method for identifying the authenticity of cucumber germplasm comprises the following steps: the method comprises the following steps: detecting the genotype of the SNP locus of the cucumber to be detected; step two: and (3) germplasm judgment of the cucumber to be detected: if the number of the genotype of the cucumber to be detected based on the 48 SNP loci and the number of the difference loci of the genotype of a certain specified germplasm in the standard cucumber germplasm based on the 48 SNP loci in the database are 0-2, the cucumber to be detected and the specified germplasm are judged to be similar germplasm; if the number of the genotype of the cucumber to be detected based on the 48 SNP loci and the number of the difference loci of the genotype of a certain specified germplasm in the standard cucumber germplasm based on the 48 SNP loci in the database are more than 2, the cucumber to be detected and the specified germplasm are judged to be different cucumber germplasms; preferably, the result of the determination is obtained from a cluster analysis.

The SNP site, or the SNP primer combination, or the SNP kit, or the DNA fingerprint database obtained by the construction method, or the detection method is applied to the following X1 or X2: x1: identifying whether the germplasm of the cucumber to be detected belongs to one of standard cucumber germplasms; x2: and identifying the specific germplasm of the cucumber to be detected as the standard cucumber germplasm.

Compared with the prior art, the invention has the following beneficial effects:

1. the invention establishes the DNA fingerprint database for identifying the authenticity of the cucumber germplasm based on high-throughput sequencing, can be used for carrying out early identification on the cucumber germplasm in tissues or organs such as seeds, seedlings, leaves and the like, practically protects the rights and interests of breeders, and provides technical support for the protection of cucumber germplasm resources.

2. The method provided by the invention can be used for identifying unknown cucumber germplasm and also can be used for identifying the authenticity of the known cucumber germplasm. The method provided by the invention has the advantages of high throughput, accuracy, low cost, simplicity in operation, manpower and material resource saving and the like, and has a very wide application prospect.

3. The cucumber germplasm SNP is mined from big data by using 110 parts of cucumber germplasm sequencing data published by a public database NCBI and 72 parts of unpublished cucumber sequencing data, candidate loci are provided for cucumber germplasm identification, specific primers of the cucumber germplasm SNP are developed by adopting an allele competitive specific PCR method, high-throughput, low-cost and automatic rapid detection is carried out, and finally the cucumber germplasm SNP genotype is obtained.

4. The reported cucumber variety identification core locus is not suitable for cucumber germplasm due to the difference of the germplasm and variety genetic characteristics of cucumber. With more than 2 different SNP loci as a threshold for identifying germplasm, only 86.81% of samples of 182 cucumber germplasm can be identified by the 32 SNP loci in the patent application with the application number of 201811634016.2, and the Pearson correlation coefficient based on genetic distance with all SNPs is 0.54, which indicates that the 32 SNPs can not effectively identify cucumber germplasm and can not represent SNPs of a whole gene. Compared with the prior art for identifying cucumber varieties based on SNP loci, the developed SNP loci for identifying cucumber germplasm can identify 182 cucumber germplasm, and have strong identification capability; the Pearson correlation coefficient based on genetic distance with the whole genome SNP is high, and the whole genome SNP can be represented; and the screening process is more complicated.

Drawings

FIG. 1 is a cluster plot of 105 cucumber germplasms tested on the 48 SNP primer sets set in example 1.

FIG. 2 is a comparison chart of the germplasm capacity of the different SNP combinations in example 1 and the 48 SNPs identified in this application. Wherein, the curve of the circular marker is the identification capability of identifying 182 cucumber germplasms by 48 SNPs in the application; the curve of the square marker is the identification capability of randomly selected 48 SNPs for identifying 182 cucumber germplasms; the curve marked by a triangle is 32 SNPs (SNP in the patent with the application number of 201811634016.2) and can identify the identification capability of 182 cucumber germplasms.

FIG. 3 is a graph showing the identification ability of the germplasm of 105 cucumber samples with 48 SNP sites in example 2.

FIG. 4 is a diagram showing the SNP typing results of 96-well sample plates for detecting 105 genotypes of cucumber germplasm in example 2 by using SNP sites numbered 1 to 12.

FIG. 5 is a diagram showing the SNP typing results of 96-well sample plates for detecting 105 genotypes of cucumber germplasm in example 2 by using SNP sites numbered 13-24.

FIG. 6 is a diagram showing the SNP typing results of 96-well sample plates for detecting 105 genotypes of cucumber germplasm in example 2 by using SNP sites numbered 25-36.

FIG. 7 is a diagram showing the SNP typing results of 96-well sample plates for detecting 105 genotypes of cucumber germplasm in example 2 by using SNP sites numbered 37-48.

Detailed Description

The definition is as follows:

and (3) cucumber germplasm authenticity: essentially refers to the real correspondence of a cucumber germplasm with its genetic background; in actual work, whether a given detected germplasm has authenticity means whether the detected germplasm conforms to a file record (such as a germplasm specification, a label and the like).

For example, the specific genomic fragment comprises but is not limited to a genomic fragment which has homology with a Chinese L ong V2 reference genomic sequence in 105 standard germplasms of the invention, and the specific genomic fragment has small mutation relative to the homologous sequence of the Chinese L ong V2 reference genomic sequence within 20 or more bases upstream and downstream of one or some of the 48 SNPs characterized by the invention, wherein the mutation is random and has no intragermplasm universality, and the SNP is considered after the mutation is ignored, so that the SNP has the intragermplasm universality.

In a first aspect, the present invention provides core SNP loci (core SNP loci are defined as a group of minimal SNP combinations capable of identifying a target germplasm and can represent genome-wide SNPs as much as possible; the core loci are distinguished from non-core loci by higher polymorphism of the core loci and capability of distinguishing the target germplasm by using minimal markers) for identifying the authenticity of cucumber germplasm, wherein the core SNP loci are selected from any 1 to 48 of the following first SNP loci to forty-eight SNP loci, as shown in Table 1:

a first SNP locus, wherein the first SNP locus is located at 215528 th chromosome 1 of a cucumber reference genome, and the nucleotide base of the first SNP locus is G or A;

a second SNP locus, wherein the second SNP locus is located at 6591765 th chromosome 1 of a cucumber reference genome, and the nucleotide base of the second SNP locus is A or T;

a third SNP locus, wherein the third SNP locus is located at 18360230 th chromosome 1 of a cucumber reference genome, and the nucleotide base of the locus is G or C;

a fourth SNP locus, wherein the fourth SNP locus is located at 19328118 th chromosome 1 of a cucumber reference genome, and the nucleotide base of the fourth SNP locus is T or C;

a fifth SNP locus, wherein the fifth SNP locus is located at 21471767 th chromosome 1 of a cucumber reference genome, and the nucleotide base of the locus is G or C;

a sixth SNP locus, wherein the sixth SNP locus is located at 26006098 th chromosome 1 of a cucumber reference genome, and the nucleotide base of the locus is A or G;

a seventh SNP locus, wherein the seventh SNP locus is located at 430448 th chromosome 2 of a cucumber reference genome, and the nucleotide base of the locus is C or T;

an eighth SNP locus, wherein the eighth SNP locus is located at 6544548 th chromosome 2 of a cucumber reference genome, and the nucleotide base of the locus is A or T;

a ninth SNP locus, wherein the ninth SNP locus is located at 7082492 th chromosome 2 of a cucumber reference genome, and the nucleotide base of the locus is A or T;

a tenth SNP locus, wherein the tenth SNP locus is located at 13016733 th chromosome 2 of a cucumber reference genome, and the nucleotide base of the locus is C or T;

an eleventh SNP locus, wherein the eleventh SNP locus is located at 19736406 th chromosome 2 of a cucumber reference genome, and the nucleotide base of the locus is C or T;

a twelfth SNP locus, wherein the twelfth SNP locus is located at 22890765 th chromosome 2 of the cucumber reference genome, and the nucleotide base of the locus is C or T;

a thirteenth SNP locus, wherein the thirteenth SNP locus is located at 3674759 th chromosome 3 of a cucumber reference genome, and the nucleotide base of the locus is G or A;

a fourteenth SNP locus, wherein the fourteenth SNP locus is located at 6715343 th chromosome 3 of a cucumber reference genome, and the nucleotide base of the locus is T or C;

a fifteenth SNP locus, wherein the fifteenth SNP locus is located at 8251132 th chromosome 3 of a cucumber reference genome, and the nucleotide base of the locus is G or A;

a sixteenth SNP locus, wherein the sixteenth SNP locus is located at 17274568 th chromosome 3 of a cucumber reference genome, and the nucleotide base of the sixteenth SNP locus is G or A;

a seventeenth SNP locus, wherein the seventeenth SNP locus is positioned at 28481554 rd chromosome of the cucumber reference genome, and the nucleotide base of the locus is C or T;

an eighteenth SNP locus, wherein the eighteenth SNP locus is located at 30072485 th chromosome of a cucumber reference genome 3, and the nucleotide base of the locus is A or G;

a nineteenth SNP locus, wherein the nineteenth SNP locus is located at 31558451 th chromosome 3 of a cucumber reference genome, and the nucleotide base of the locus is A or G;

a twentieth SNP locus, wherein the twentieth SNP locus is located at 1284842 th chromosome 4 of the cucumber reference genome, and the nucleotide base of the locus is G or T;

a twenty-first SNP locus, wherein the twenty-first SNP locus is located at 14339957 th chromosome 4 of a cucumber reference genome, and the nucleotide base of the locus is A or G;

a twenty-second SNP locus, wherein the twenty-second SNP locus is located at 16172836 th chromosome 4 of a cucumber reference genome, and the nucleotide base of the locus is A or G;

a twenty-third SNP locus, wherein the twenty-third SNP locus is located at 20428654 th chromosome 4 of a cucumber reference genome, and the nucleotide base of the locus is G or A;

a twenty-fourth SNP locus, wherein the twenty-fourth SNP locus is located at 22774845 th chromosome 4 of a cucumber reference genome, and the nucleotide base of the locus is G or A;

a twenty-fifth SNP locus, wherein the twenty-fifth SNP locus is located at 1045863 th chromosome 5 of a cucumber reference genome, and the nucleotide base of the locus is T or G;

a twenty-sixth SNP locus, wherein the twenty-sixth SNP locus is located at 4850607 th chromosome 5 of a cucumber reference genome, and the nucleotide base of the locus is G or A;

a twenty-seventh SNP locus, wherein the twenty-seventh SNP locus is located at 5787330 th chromosome 5 of a cucumber reference genome, and the nucleotide base of the locus is T or A;

a twenty-eighth SNP locus, wherein the twenty-eighth SNP locus is located at 11869197 th chromosome 5 of a cucumber reference genome, and the nucleotide base of the locus is G or C;

a twenty-ninth SNP locus, wherein the twenty-ninth SNP locus is located at 15556517 th chromosome 5 of a cucumber reference genome, and the nucleotide base of the locus is T or G;

a thirtieth SNP locus, wherein the thirtieth SNP locus is located at 23713010 th chromosome 5 of a cucumber reference genome, and the nucleotide base of the locus is T or C;

a thirty-first SNP locus, wherein the thirty-first SNP locus is located at 24681408 th chromosome 5 of a cucumber reference genome, and the nucleotide base of the locus is C or A;

a thirty-second SNP locus, wherein the thirty-second SNP locus is located at 437035 th chromosome of the cucumber reference genome 6, and the nucleotide base of the locus is G or C;

a thirty-third SNP locus, wherein the thirty-third SNP locus is located at 6311543 th chromosome 6 of a cucumber reference genome, and the nucleotide base of the locus is T or G;

a thirty-fourth SNP locus, wherein the thirty-fourth SNP locus is located at 7644442 th chromosome 6 of a cucumber reference genome, and the nucleotide base of the locus is T or C;

a thirty-fifth SNP locus, wherein the thirty-fifth SNP locus is located at 8554160 th chromosome of a cucumber reference genome 6, and the nucleotide base of the locus is C or T;

a thirty-sixth SNP locus, wherein the thirty-sixth SNP locus is located at 9796426 th chromosome of the cucumber reference genome 6, and the nucleotide base of the locus is G or A;

a thirty-seventh SNP locus, wherein the thirty-seventh SNP locus is located at 11577012 th chromosome 6 of a cucumber reference genome, and the nucleotide base of the locus is T or G;

a thirty-eighth SNP locus, wherein the thirty-eighth SNP locus is located at 13951516 th chromosome of a cucumber reference genome 6, and the nucleotide base of the locus is T or A;

a thirty-ninth SNP locus, wherein the thirty-ninth SNP locus is located at 18948506 th chromosome 6 of a cucumber reference genome, and the nucleotide base of the locus is G or A;

a fortieth SNP locus, wherein the second SNP locus is located at 24605064 th chromosome of a 6 th chromosome of a cucumber reference genome, and the nucleotide base of the locus is A or T;

a forty-first SNP locus, wherein the fourth eleventh SNP locus is located at 4474790 th chromosome 7 of a cucumber reference genome, and the nucleotide base of the locus is C or T;

a forty-second SNP locus, wherein the fourth twelve SNP locus is located at 6777921 th chromosome 7 of a cucumber reference genome, and the nucleotide base of the locus is C or A;

a forty-third SNP locus, wherein the forty-fourth SNP locus is located at 7779724 th chromosome 7 of a cucumber reference genome, and the nucleotide base of the locus is G or A;

a forty-fourth SNP locus, wherein the forty-fourth SNP locus is located at 9945305 th chromosome 7 of a cucumber reference genome, and the nucleotide base of the locus is T or C;

a forty-fifth SNP locus, wherein the forty-fifth SNP locus is located at 10495910 th chromosome 7 of a cucumber reference genome, and the nucleotide base of the locus is A or G;

a forty-sixth SNP locus, wherein the forty-sixth SNP locus is located at 11918569 th chromosome 7 of a cucumber reference genome, and the nucleotide base of the locus is C or T;

a forty-seventh SNP locus, wherein the forty-seventh SNP locus is located at 12580918 th chromosome 7 of a cucumber reference genome, and the nucleotide base of the locus is G or A;

a forty-eighth SNP locus, wherein the forty-eighth SNP locus is located at 14166928 th chromosome 7 of a cucumber reference genome, and the nucleotide base of the locus is A or G.

The cucumber reference genome is Chinese L ong V2.

The sequences of the first SNP locus and bases at the upstream and downstream are SEQ ID NO: 145 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 145 greater than or equal to 95%, 96%, 97%, 98% or 99% identity;

the sequences of the second SNP locus and bases at the upstream and downstream of the second SNP locus are SEQ ID NO: 146 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 146, greater than or equal to 95%, 96%, 97%, 98%, or 99%;

the sequences of the third SNP locus and bases at the upstream and downstream are SEQ ID NO: 147 or an idioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 147 is greater than or equal to 95%, 96%, 97%, 98% or 99% identical;

the fourth SNP locus and the sequences of the upstream and downstream bases thereof are SEQ ID NO: 148 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 148 greater than or equal to 95%, 96%, 97%, 98%, or 99%;

the fifth SNP locus and the sequences of the upstream and downstream bases thereof are SEQ ID NO: 149 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 149 is greater than or equal to 95%, 96%, 97%, 98% or 99%;

the sequences of the sixth SNP locus and the upstream and downstream bases thereof are SEQ ID NO: 150 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 150 is greater than or equal to 95%, 96%, 97%, 98%, or 99%;

the seventh SNP locus and the sequences of bases on the seventh SNP locus and upstream and downstream thereof are SEQ ID NO: 151 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 151 is greater than or equal to 95%, 96%, 97%, 98%, or 99% identical;

the sequences of the eighth SNP locus and bases at the upstream and downstream are SEQ ID NO: 152 or an interidioplasmically homologous genomic fragment thereof, more preferably to SEQ ID NO: 152 is greater than or equal to 95%, 96%, 97%, 98%, or 99% identical;

the ninth SNP site and the sequences of the upstream and downstream bases thereof are SEQ ID NO: 153 or an idioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 153 by greater than or equal to 95%, 96%, 97%, 98% or 99%;

the tenth SNP site and the sequences of the upstream and downstream bases thereof are SEQ ID NO: 154 or an interidioplasmically homologous genomic fragment thereof, more preferably to SEQ ID NO: 154 greater than or equal to 95%, 96%, 97%, 98%, or 99%;

the sequence of the eleventh SNP site and bases on the eleventh SNP site is SEQ ID NO: 155 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 155 greater than or equal to 95%, 96%, 97%, 98%, or 99%;

the sequence of the twelfth SNP site and the upstream and downstream bases thereof is SEQ ID NO: 156 or an idioplasmically homologous genomic fragment thereof, more preferably to SEQ ID NO: 156 is greater than or equal to 95%, 96%, 97%, 98%, or 99%;

the thirteenth SNP site and the sequences of the upstream and downstream bases thereof are SEQ ID NO: 157 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 157 greater than or equal to 95%, 96%, 97%, 98%, or 99%;

the sequence of the fourteenth SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 158 or an interidioplasmically homologous genomic fragment thereof, more preferably to SEQ ID NO: 158 is greater than or equal to 95%, 96%, 97%, 98% or 99% identical;

the sequence of the fifteenth SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 159 or an interidioplasmically homologous genomic fragment thereof, more preferably a fragment of SEQ ID NO: 159 is greater than or equal to 95%, 96%, 97%, 98% or 99%;

the sequence of the sixteenth SNP locus and bases on the sixteenth SNP locus is SEQ ID NO: 160 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 160, greater than or equal to 95%, 96%, 97%, 98%, or 99%;

the sequence of the seventeenth SNP site and the upstream and downstream bases thereof is SEQ ID NO: 161 or an interidioplasmic homologous genomic fragment thereof, more preferably identical to SEQ ID NO: 161 is greater than or equal to 95%, 96%, 97%, 98% or 99% identical;

the sequence of the eighteenth SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 162 or an idioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 162, greater than or equal to 95%, 96%, 97%, 98%, or 99%;

the nineteenth SNP site and the sequences of bases on the nineteenth SNP site are SEQ ID NO: 163 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 163, greater than or equal to 95%, 96%, 97%, 98%, or 99%;

the twenty-second SNP site and the sequences of bases on the twenty-second SNP site are SEQ ID NO: 164 or an interidioplasmically homologous genomic fragment thereof, more preferably to the sequence set forth in SEQ ID NO: 164 is greater than or equal to 95%, 96%, 97%, 98% or 99% identical;

the twenty-first SNP locus and the sequences of bases on the twenty-first SNP locus and bases on the twenty-first SNP locus are SEQ ID NO: 165 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 165 greater than or equal to 95%, 96%, 97%, 98% or 99% identity;

the sequence of the second twelve SNP locus and the base sequences of the second twelve SNP locus are SEQ ID NO: 166 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 166 is greater than or equal to 95%, 96%, 97%, 98%, or 99% identical;

the sequence of the twenty-third SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 167 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 167 nucleotide sequence identity of greater than or equal to 95%, 96%, 97%, 98%, or 99%;

the sequence of the twenty-fourth SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 168 or an interidioplasmic homologous genomic fragment thereof, more preferably to the sequence set forth in SEQ ID NO: 168, greater than or equal to 95%, 96%, 97%, 98%, or 99%;

the twenty-fifth SNP locus and the sequences of bases on the twenty-fifth SNP locus and the upstream and downstream of the twenty-fifth SNP locus are SEQ ID NO: 169 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 169 greater than or equal to 95%, 96%, 97%, 98%, or 99% identity in the nucleotide sequence;

the twenty-sixth SNP locus and the sequences of bases on the twenty-sixth SNP locus and bases on the twenty-sixth SNP locus are SEQ ID NO: 170 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 170, greater than or equal to 95%, 96%, 97%, 98%, or 99%;

the twenty-seventh SNP locus and the sequences of bases on the twenty-seventh SNP locus and the upstream and downstream of the twenty-seventh SNP locus are SEQ ID NO: 171 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 171 is greater than or equal to 95%, 96%, 97%, 98%, or 99% identical;

the sequences of the twenty-eight SNP locus and the upstream and downstream bases thereof are SEQ ID NO: 172 or an interidioplasmically homologous genomic fragment thereof, more preferably identical to SEQ ID NO: 172 is greater than or equal to 95%, 96%, 97%, 98%, or 99% identical;

the twenty-ninth SNP locus and the sequences of bases on the twenty-ninth SNP locus are SEQ ID NO: 173 or an interidioplasmically homologous genomic fragment thereof, more preferably to SEQ ID NO: 173, greater than or equal to 95%, 96%, 97%, 98%, or 99%;

the thirty-third SNP site and the sequences of the upstream and downstream bases thereof are SEQ ID NO: 174 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 174, or greater than or equal to 95%, 96%, 97%, 98%, or 99%;

the sequence of the thirty-first SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 175 or an idioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 175 greater than or equal to 95%, 96%, 97%, 98%, or 99%;

the sequences of the third twelve SNP loci and bases at the upper and lower ends of the third twelve SNP loci are SEQ ID NO: 176 or an interidioplasmic homologous genomic fragment thereof, more preferably identical to SEQ ID NO: 176, greater than or equal to 95%, 96%, 97%, 98%, or 99%;

the sequence of the thirteenth SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 177 or an idioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 177 greater than or equal to 95%, 96%, 97%, 98%, or 99% identity;

the sequence of the thirty-fourth SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 178 or an idioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 178 greater than or equal to 95%, 96%, 97%, 98%, or 99%;

the sequence of the fifteenth SNP locus and the bases at the upstream and downstream of the fifteenth SNP locus is SEQ ID NO: 179 or an interidioplasmically homologous genomic fragment thereof, more preferably to SEQ ID NO: 179 greater than or equal to 95%, 96%, 97%, 98% or 99% identity;

the sequences of the sixteenth SNP locus and bases on the sixteenth SNP locus are SEQ ID NO: 180 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 180 greater than or equal to 95%, 96%, 97%, 98% or 99%;

the sequences of the seventeenth SNP site and the upstream and downstream bases thereof are SEQ ID NO: 181 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 181 greater than or equal to 95%, 96%, 97%, 98%, or 99% identity;

the sequences of the third eighteen SNP locus and the upstream and downstream bases thereof are SEQ ID NO: 182 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 182 is greater than or equal to 95%, 96%, 97%, 98%, or 99% identical;

the sequence of the thirty-ninth SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 183 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 183 greater than or equal to 95%, 96%, 97%, 98% or 99% identity;

the sequence of the fortieth SNP locus and bases on the fortieth SNP locus are SEQ ID NO: 184 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 184 is greater than or equal to 95%, 96%, 97%, 98% or 99% identical;

the sequence of the fourth eleventh SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 185 or an idioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 185 is greater than or equal to 95%, 96%, 97%, 98% or 99%;

the sequence of the fourth twelve SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 186 or an idioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 186 nucleotide sequence identity of greater than or equal to 95%, 96%, 97%, 98%, or 99%;

the sequence of the fourth and the thirteenth SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 187 or an interidioplasmic homologous genomic fragment thereof, more preferably to the sequence set forth in SEQ ID NO: 187 greater than or equal to 95%, 96%, 97%, 98% or 99% identity;

the sequence of the fourteenth SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 188 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 188 is greater than or equal to 95%, 96%, 97%, 98%, or 99%;

the sequence of the fifteenth SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 189 or an intergenic homologous genomic fragment thereof, more preferably to SEQ ID NO: 189 is greater than or equal to 95%, 96%, 97%, 98% or 99% identical;

the sequence of the sixteenth SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 190 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 190 is greater than or equal to 95%, 96%, 97%, 98% or 99%;

the sequence of the fourth seventeen SNP site and the upstream and downstream bases thereof is SEQ ID NO: 191 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 191, greater than or equal to 95%, 96%, 97%, 98% or 99%;

the sequence of the fourth eighteenth SNP locus and the upstream and downstream bases thereof is SEQ ID NO: 192 or an interidioplasmic homologous genomic fragment thereof, more preferably to SEQ ID NO: 192 of greater than or equal to 95%, 96%, 97%, 98% or 99%.

In a second aspect, the present invention provides a combination of core SNP primer sets for identifying the genuineness of cucumber germplasm, comprising:

a first SNP primer set for amplifying the first SNP site; a second SNP primer set for amplifying the second SNP site; a third SNP primer set for amplifying the third SNP site; a fourth SNP primer set for amplifying the fourth SNP site; a fifth SNP primer set for amplifying the fifth SNP site; a sixth SNP primer set for amplifying the sixth SNP site; a seventh SNP primer set for amplifying the seventh SNP site; an eighth SNP primer set for amplifying the eighth SNP site; a ninth SNP primer set for amplifying the ninth SNP site; a tenth SNP primer set for amplifying the tenth SNP site; an eleventh SNP primer set for amplifying the eleventh SNP site; a twelfth SNP primer set for amplifying the twelfth SNP site; a thirteenth SNP primer set for amplifying the thirteenth SNP site; a fourteenth SNP primer set for amplifying the fourteenth SNP site; a fifteenth SNP primer set for amplifying the fifteenth SNP site; a sixteenth SNP primer set for amplifying the sixteenth SNP site; a seventeenth SNP primer set for amplifying the seventeenth SNP site; an eighteenth SNP primer set for amplifying the eighteenth SNP site; a nineteenth SNP primer set for amplifying the nineteenth SNP site; a twentieth SNP primer set for amplifying the twentieth SNP site; a twenty-first SNP primer set for amplifying the twenty-first SNP site; a second twelve SNP primer set for amplifying the second twelve SNP sites; a twenty-third SNP primer set for amplifying the twenty-third SNP site; a twenty-fourth SNP primer set for amplifying the twenty-fourth SNP site; a twenty-fifth SNP primer set for amplifying the twenty-fifth SNP site; a twenty-sixth SNP primer set for amplifying the twenty-sixth SNP site; a twenty-seventh SNP primer set for amplifying the twenty-seventh SNP site; a second eighteen SNP primer set for amplifying the second eighteen SNP site; a twenty-ninth SNP primer set for amplifying the twenty-ninth SNP site; a thirtieth SNP primer set for amplifying the thirtieth SNP site; a thirty-first SNP primer set for amplifying the thirty-first SNP site; a third twelve SNP primer set for amplifying the third twelve SNP sites; a thirteenth SNP primer set for amplifying the thirteenth SNP site; a thirty-fourth SNP primer set for amplifying the thirty-fourth SNP site; a fifteenth SNP primer set for amplifying the fifteenth SNP site; a sixteenth SNP primer set for amplifying the sixteenth SNP site; a third seventeen SNP primer set for amplifying the third seventeen SNP site; a third eighteen SNP primer set for amplifying the third eighteen SNP site; a thirty-ninth SNP primer set for amplifying the thirty-ninth SNP site; a fortieth SNP primer set for amplifying the fortieth SNP site; a fourth eleventh SNP primer set for amplifying the fourth eleventh SNP site; a fourth twelve SNP primer set for amplifying the fourth twelve SNP sites; a thirteenth SNP primer set for amplifying the thirteenth SNP site; a fourteenth SNP primer set for amplifying the fourteenth SNP site; a fifteenth SNP primer set for amplifying the fifteenth SNP site; a sixteenth SNP primer set for amplifying the sixteenth SNP site; a seventeenth SNP primer set for amplifying the seventeenth SNP site; a forty-eighth SNP primer set for amplifying the forty-eighth SNP site.

In some embodiments, the first SNP primer set, including the specific portion of the first upstream primer (F1), the specific portion of the second upstream primer (F2) of the first SNP primer set, the downstream primer (R) of the first SNP primer set, are identical to SEQ ID NOs: 1. SEQ ID NO: 2. SEQ ID NO: 3 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

the second SNP primer set, which includes a specific portion of the first upstream primer (F1) of the second SNP primer set, a specific portion of the second upstream primer (F2) of the second SNP primer set, and a downstream primer (R) of the second SNP primer set, are identical to SEQ ID NO: 4. SEQ ID NO: 5. SEQ ID NO: 6 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

the third SNP primer set, which includes a specific portion of the first upstream primer (F1) of the third SNP primer set, a specific portion of the second upstream primer (F2) of the third SNP primer set, and a downstream primer (R) of the third SNP primer set, are identical to SEQ ID NO: 7. SEQ ID NO: 8. SEQ ID NO: 9 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

the fourth SNP primer set, including the specific portion of the first upstream primer (F1), the specific portion of the second upstream primer (F2), and the downstream primer (R), of the fourth SNP primer set, are linked to SEQ ID NO: 10. SEQ ID NO: 11. SEQ ID NO: 12 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

the fifth SNP primer set, including the specific portion of the first upstream primer (F1), the specific portion of the second upstream primer (F2), and the downstream primer (R), of the fifth SNP primer set, are linked to SEQ ID NO: 13. SEQ ID NO: 14. SEQ ID NO: 15, is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98%, or 99%, preferably 100%;

the sixth SNP primer set, including the specific portion of the first upstream primer (F1) of the sixth SNP primer set, the specific portion of the second upstream primer (F2) of the sixth SNP primer set, and the downstream primer (R) of the sixth SNP primer set, are linked to SEQ ID NOs: 16. SEQ ID NO: 17. SEQ ID NO: 18, is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98%, or 99%, preferably 100%;

the seventh SNP primer set, including the specific portion of the first upstream primer (F1) of the seventh SNP primer set, the specific portion of the second upstream primer (F2) of the seventh SNP primer set, and the downstream primer (R) of the seventh SNP primer set, are linked to SEQ ID NOs: 19. SEQ ID NO: 20. SEQ ID NO: 21 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

the eighth SNP primer set, including the specific portion of the first upstream primer (F1) of the eighth SNP primer set, the specific portion of the second upstream primer (F2) of the eighth SNP primer set, and the downstream primer (R) of the eighth SNP primer set, are linked to SEQ ID NOs: 22. SEQ ID NO: 23. SEQ ID NO: 24 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

the ninth SNP primer set, including the specific portion of the first upstream primer (F1) of the ninth SNP primer set, the specific portion of the second upstream primer (F2) of the ninth SNP primer set, and the downstream primer (R) of the ninth SNP primer set, are linked to SEQ ID NOs: 25. SEQ ID NO: 26. SEQ ID NO: 27 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

the tenth SNP primer set, including the specific portion of the first upstream primer (F1) of the tenth SNP primer set, the specific portion of the second upstream primer (F2) of the tenth SNP primer set, and the downstream primer (R) of the tenth SNP primer set, are linked to SEQ ID NOs: 28. SEQ ID NO: 29. SEQ ID NO: 30 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

the eleventh SNP primer set, including the specific portion of the first upstream primer (F1) of the eleventh SNP primer set, the specific portion of the second upstream primer (F2) of the eleventh SNP primer set, and the downstream primer (R) of the eleventh SNP primer set, are linked to SEQ ID NOs: 31. SEQ ID NO: 32. SEQ ID NO: 33, is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98%, or 99%, preferably 100%;

the twelfth SNP primer set, which includes a specific portion of the first upstream primer (F1) of the twelfth SNP primer set, a specific portion of the second upstream primer (F2) of the twelfth SNP primer set, and a downstream primer (R) of the twelfth SNP primer set, is identical to the sequence set shown in SEQ ID NO: 34. SEQ ID NO: 35. SEQ ID NO: 36 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

the thirteenth SNP primer set, including the specific portion of the first upstream primer (F1) of the thirteenth SNP primer set, the specific portion of the second upstream primer (F2) of the thirteenth SNP primer set, the downstream primer (R) of the thirteenth SNP primer set, is identical to SEQ ID NO: 37. SEQ ID NO: 38. SEQ ID NO: 39, greater than or equal to 85%, 90%, 95%, 96%, 97%, 98%, or 99%, preferably 100%;

the fourteenth SNP primer set, which includes a specific portion of the first upstream primer (F1) of the fourteenth SNP primer set, a specific portion of the second upstream primer (F2) of the fourteenth SNP primer set, and a downstream primer (R) of the fourteenth SNP primer set, is identical to SEQ ID NO: 40. SEQ ID NO: 41. SEQ ID NO: 42 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

the fifteenth SNP primer set, including the specific portion of the first upstream primer (F1), the specific portion of the second upstream primer (F2), and the downstream primer (R), of the fifteenth SNP primer set, are linked to SEQ ID NOs: 43. SEQ ID NO: 44. SEQ ID NO: 45 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

the sixteenth SNP primer set, including the specific portion of the first upstream primer (F1) of the sixteenth SNP primer set, the specific portion of the second upstream primer (F2) of the sixteenth SNP primer set, and the downstream primer (R) of the sixteenth SNP primer set, are linked to SEQ ID NOs: 46. SEQ ID NO: 47. SEQ ID NO: 48, is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98%, or 99%, preferably 100%;

the seventeenth SNP primer set, including the specific portion of the first upstream primer (F1) of the seventeenth SNP primer set, the specific portion of the second upstream primer (F2) of the seventeenth SNP primer set, and the downstream primer (R) of the seventeenth SNP primer set, are linked to SEQ ID NOs: 49. SEQ ID NO: 50. SEQ ID NO: 51, is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98%, or 99%, preferably 100%;

the eighteenth SNP primer set, including the specific portion of the first upstream primer (F1) of the eighteenth SNP primer set, the specific portion of the second upstream primer (F2) of the eighteenth SNP primer set, and the downstream primer (R) of the eighteenth SNP primer set, are linked to SEQ ID NOs: 52. SEQ ID NO: 53. SEQ ID NO: 54 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

the nineteenth SNP primer set, including the specific portion of the first upstream primer (F1) of the nineteenth SNP primer set, the specific portion of the second upstream primer (F2) of the nineteenth SNP primer set, and the downstream primer (R) of the nineteenth SNP primer set, are identical to SEQ ID NOs: 55. SEQ ID NO: 56. SEQ ID NO: 57 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

the twentieth SNP primer set, including the specific portion of the first upstream primer (F1) of the twentieth SNP primer set, the specific portion of the second upstream primer (F2) of the twentieth SNP primer set, and the downstream primer (R) of the twentieth SNP primer set, are linked to SEQ ID NOs: 58. SEQ ID NO: 59. SEQ ID NO: 60, preferably 100%, or greater than 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity;

the twenty-first SNP primer set including a specific portion of the first upstream primer (F1) of the twenty-first SNP primer set, a specific portion of the second upstream primer (F2) of the twenty-first SNP primer set, a downstream primer (R) of the twenty-first SNP primer set, and a sequence listing corresponding to SEQ ID NOs: 61. SEQ ID NO: 62. SEQ ID NO: 63 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

the second twelve SNP primer set including the specific portion of the first upstream primer (F1), the specific portion of the second upstream primer (F2) of the second twelve SNP primer set, the downstream primer (R) of the second twelve SNP primer set, and the sequence numbers of SEQ ID NOs: 64. SEQ ID NO: 65. SEQ ID NO: 66 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

the thirteenth SNP primer set including a specific portion of the first upstream primer (F1) of the thirteenth SNP primer set, a specific portion of the second upstream primer (F2) of the thirteenth SNP primer set, and a downstream primer (R) of the thirteenth SNP primer set, are identical to SEQ ID NOs: 67. SEQ ID NO: 68. SEQ ID NO: 69 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

the twenty-fourth SNP primer set, including the specific portion of the first upstream primer (F1), the specific portion of the second upstream primer (F2), and the downstream primer (R), of the twenty-fourth SNP primer set, are identical to SEQ ID NOs: 70. SEQ ID NO: 71. SEQ ID NO: 72 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

the twenty-fifth SNP primer set, including the specific portion of the first upstream primer (F1), the specific portion of the second upstream primer (F2), and the downstream primer (R), of the twenty-fifth SNP primer set, are identical to SEQ ID NOs: 73. SEQ ID NO: 74. SEQ ID NO: 75 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

the twenty-sixth SNP primer set including the specific portion of the first upstream primer (F1), the specific portion of the second upstream primer (F2) and the downstream primer (R) of the twenty-sixth SNP primer set, respectively, have the same sequence as SEQ ID NO: 76. SEQ ID NO: 77. SEQ ID NO: 78, is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98%, or 99%, preferably 100%;

the twenty-seventh SNP primer set, including the specific portion of the first upstream primer (F1), the specific portion of the second upstream primer (F2), and the downstream primer (R), of the twenty-seventh SNP primer set, are identical to SEQ ID NOs: 79. SEQ ID NO: 80. SEQ ID NO: 81 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

the second eighteen SNP primer set, including the specific portion of the first upstream primer (F1), the specific portion of the second upstream primer (F2), the downstream primer (R), of the second eighteen SNP primer set, is identical to SEQ ID NO: 82. SEQ ID NO: 83. SEQ ID NO: 84 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

the twenty-ninth SNP primer set including the specific portion of the first upstream primer (F1), the specific portion of the second upstream primer (F2) and the downstream primer (R) of the twenty-ninth SNP primer set, respectively, have the same sequence as SEQ ID NO: 85. SEQ ID NO: 86. SEQ ID NO: 87 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

the thirtieth SNP primer set comprises a specific part of the first upstream primer (F1) of the thirtieth SNP primer set, a specific part of the second upstream primer (F2) of the thirtieth SNP primer set and a downstream primer (R) of the thirtieth SNP primer set, which are respectively connected with the sequence numbers of SEQ ID NOs: 88. SEQ ID NO: 89. SEQ ID NO: 90 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

the thirty-first SNP primer set, including the specific portion of the first upstream primer (F1), the specific portion of the second upstream primer (F2), and the downstream primer (R), of the thirty-first SNP primer set, are identical to SEQ ID NOs: 91. SEQ ID NO: 92. SEQ ID NO: 93 greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

the third twelve SNP primer set including the specific portion of the first upstream primer (F1), the specific portion of the second upstream primer (F2) and the downstream primer (R) of the third twelve SNP primer set, respectively, are identical to SEQ ID NOs: 94. SEQ ID NO: 95. SEQ ID NO: 96 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

the thirteenth SNP primer set including a specific portion of the first upstream primer (F1), a specific portion of the second upstream primer (F2), and a downstream primer (R) of the thirteenth SNP primer set, respectively, overlap with SEQ ID NOs: 97. SEQ ID NO: 98. SEQ ID NO: 99, and preferably 100%, or greater than 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity;

the thirty-fourth SNP primer set, which comprises a specific part of the first upstream primer (F1) of the thirty-fourth SNP primer set, a specific part of the second upstream primer (F2) of the thirty-fourth SNP primer set, and a downstream primer (R) of the thirty-fourth SNP primer set, is respectively identical to the sequence of SEQ ID NO: 100. SEQ ID NO: 101. SEQ ID NO: 102 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

the fifteenth SNP primer set including the specific portion of the first upstream primer (F1), the specific portion of the second upstream primer (F2), and the downstream primer (R) of the fifteenth SNP primer set, respectively, are identical to SEQ ID NOs: 103. SEQ ID NO: 104. SEQ ID NO: 105 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

the third sixteen SNP primer set including the specific portion of the first upstream primer (F1), the specific portion of the second upstream primer (F2), and the downstream primer (R) of the third sixteen SNP primer set, respectively, have the same sequence as SEQ ID NOs: 106. SEQ ID NO: 107. SEQ ID NO: 108 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

the third seventeen SNP primer set, including the specific portion of the first upstream primer (F1), the specific portion of the second upstream primer (F2), and the downstream primer (R), of the third seventeen SNP primer set, are identical to SEQ ID NOs: 109. SEQ ID NO: 110. SEQ ID NO: 111 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

the thirty-eighth SNP primer set including a specific portion of the first upstream primer (F1), a specific portion of the second upstream primer (F2) of the thirty-eighth SNP primer set, and a downstream primer (R) of the thirty-eighth SNP primer set, respectively, have the same sequence as SEQ ID NOs: 112. SEQ ID NO: 113. SEQ ID NO: 114 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

the third nineteenth SNP primer set, including the specific portion of the first upstream primer (F1), the specific portion of the second upstream primer (F2), and the downstream primer (R), of the third nineteenth SNP primer set, are identical to SEQ ID NOs: 115. SEQ ID NO: 116. SEQ ID NO: 117 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

the fortieth SNP primer set including the specific portion of the first upstream primer (F1) of the fortieth SNP primer set, the specific portion of the second upstream primer (F2) of the fortieth SNP primer set, and the downstream primer (R) of the fortieth SNP primer set, are respectively identical to SEQ ID NOs: 118. SEQ ID NO: 119. SEQ ID NO: 120, is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98%, or 99%, preferably 100%;

the fourth eleventh SNP primer set, including the specific portion of the first upstream primer (F1), the specific portion of the second upstream primer (F2), and the downstream primer (R), of the fourth eleventh SNP primer set, are identical to SEQ ID NOs: 121. SEQ ID NO: 122. SEQ ID NO: 123 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

the fourth twelve SNP primer set including the specific portion of the first upstream primer (F1), the specific portion of the second upstream primer (F2) of the fourth twelve SNP primer set, and the downstream primer (R) of the fourth twelve SNP primer set, are identical to SEQ ID NOs: 124. SEQ ID NO: 125. SEQ ID NO: 126, is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98%, or 99%, preferably 100%;

the thirteenth SNP primer set including a specific portion of the first upstream primer (F1) of the thirteenth SNP primer set, a specific portion of the second upstream primer (F2) of the thirteenth SNP primer set, and a downstream primer (R) of the thirteenth SNP primer set, are identical to SEQ ID NOs: 127. SEQ ID NO: 128. SEQ ID NO: 129 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

the fourteenth SNP primer set, which includes a specific portion of the first upstream primer (F1) of the fourteenth SNP primer set, a specific portion of the second upstream primer (F2) of the fourteenth SNP primer set, and a downstream primer (R) of the fourteenth SNP primer set, is identical to SEQ ID NO: 130. SEQ ID NO: 131. SEQ ID NO: 132 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

the fifteenth SNP primer set, including the specific portion of the first upstream primer (F1), the specific portion of the second upstream primer (F2), and the downstream primer (R), of the fifteenth SNP primer set, are linked to SEQ ID NOs: 133. SEQ ID NO: 134. SEQ ID NO: 135 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

the sixteenth SNP primer set, including the specific portion of the first upstream primer (F1), the specific portion of the second upstream primer (F2), and the downstream primer (R), of the sixteenth SNP primer set, are identical to SEQ ID NOs: 136. SEQ ID NO: 137. SEQ ID NO: 138 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%;

the fourth seventeen SNP primer set, including the specific portion of the first upstream primer (F1) of the fourth seventeen SNP primer set, the specific portion of the second upstream primer (F2) of the fourth seventeen SNP primer set, the downstream primer (R) of the fourth seventeen SNP primer set, is identical to SEQ ID NO: 139. SEQ ID NO: 140. SEQ ID NO: 141, greater than or equal to 85%, 90%, 95%, 96%, 97%, 98%, or 99%, preferably 100%;

the fourth eighteenth SNP primer set, including the specific portion of the first upstream primer (F1) of the fourth eighteenth SNP primer set, the specific portion of the second upstream primer (F2) of the fourth eighteenth SNP primer set, the downstream primer (R) of the fourth eighteenth SNP primer set, is identical to SEQ ID NO: 142. SEQ ID NO: 143. SEQ ID NO: 144 is greater than or equal to 85%, 90%, 95%, 96%, 97%, 98% or 99%, preferably 100%.

Preferably, one primer of each of said primer pairs is linked to a fluorescent molecule, more preferably said fluorescent molecule is selected from the group consisting of ROX, TAMRA, FAM, HEX.

In a preferred embodiment, the SNP primer combination is selected from one or more of the primer sets 01 to 48; the DNA sequence information of the primer groups 01-48 is shown in a sequence table SEQ ID: 1-144, see table 2.

In the primer group, the 5' end of the upstream primer can be provided with a fluorescent label sequence for fluorescent PCR detection, the first upstream primer and the second upstream primer in each group of primers are connected with different fluorescent molecules, and more preferably, the fluorescent molecules are selected from ROX, TAMRA, FAM and HEX;

for example, the fluorescence signal of FAM fluorescent tag sequence is blue, and the fluorescence signal of HEX fluorescent tag sequence is red.

In a third aspect, the present invention provides a core SNP kit for identifying the genuineness of cucumber germplasm, the SNP reagent being formulated as a competitive allele-specific PCR reaction system, preferably comprising:

in the SNP primer sets, the concentration ratio of the first upstream primer, the second upstream primer and the downstream primer of each primer set in the system is 2:2: 5;

the reagents, consumables and apparatus in the reaction system were all provided by L GC company, including reagent amounts, usage, and overall experimental procedures, were performed according to the operating manual KASP user guide and manual (www.lgcgenomics.com) of L GC company, the KASPar reaction was performed in 384 well plates (Part No. KBS-0750-001) or 96 well plates (Part No. KBS-0751-001), and the reaction system was 3ul or 1ul, as shown in the following table.

Table: KASP reaction system of 384-well plate or 96-well plate

The preparation method of the kit comprises the step of packaging each primer in any one primer group separately.

In a fourth aspect, the present invention provides the above cucumber germplasm DNA fingerprint database based on the core SNP markers, wherein the DNA fingerprint database comprises: and (3) the genotype of the core SNP locus of the standard cucumber germplasm.

The standard cucumber germplasm comprises 105 cucumber germplasms selected from the following:

beijing lesser spiny melon, CM8537, pacific pine leaf three, green tangerine peel eight-crotch, Roqing 7314-2-6-1-1, Liaotong dense thorn, yellow iron non-spiny melon, bvrc, Cucumis sativus, leaf tribe, Liaoyang leaf three, Xintai dense thorn, bvrc, bvrc, bvrc, Qingdao autumn leaf three, Kaga Fushinari, Sagami Hanpaku Fushinari Kyuri, anonga Suyo Kyuri, uda, Sakata Natsunsang, Honshu Aibai, Sekino No.2(Ochiai No.2), CGN19828, bvrc, Cucumis sativus, autumn cucumber, dew, bvlon, bvrc, bvrc, bvrc, Sichuan white melon, bvre, Bvre, Purch No.2, Kirch No.2, Purch No. 4, Purch-7, Purch No. 4, Purch No.2, Purch No. 4, Purch-7, Purch No. 4, Purch No. 4, Kvkura, Kyu.

In a fifth aspect, the invention provides a method for constructing the cucumber germplasm DNA fingerprint database, which comprises the following steps:

s1: KASP reaction: carrying out competitive allele specificity PCR amplification reaction on the standard cucumber germplasm by adopting the reaction system to obtain a PCR reaction product;

the PCR reaction program is: pre-denaturation at 94 ℃ for 15 min; denaturation at 94 ℃ for 20s, denaturation at 61-55 ℃ (touch down program is selected, reduction of 0.6 ℃ per cycle) is carried out, 1min is carried out, and amplification is carried out for 10 cycles; denaturation at 94 ℃ for 20s, renaturation at 55 ℃ and extension for 1min, and amplification is continued for 26 cycles.

S2: SNP locus genotype acquisition: and detecting the PCR reaction product to obtain the genotype of the SNP locus.

The above detection method may be selected from: for fluorescent signal detection, direct sequencing and restriction enzyme digestion are carried out.

In a sixth aspect, the invention provides an authenticity detection method for identifying cucumber germplasm, comprising the following steps:

s1, detecting the genotype of the SNP locus of the cucumber to be detected:

respectively taking the genome DNA of cucumber to be detected as a template, and respectively adopting the primer groups in the SNP primer combination to carry out competitive allele specific PCR amplification reaction to obtain PCR amplification products;

s2, germplasm judgment of the cucumber to be detected: comparing the genotype of the SNP locus of the cucumber to be detected with the DNA fingerprint database, obtaining a result through cluster analysis and judging, wherein the judgment standard is as follows:

if the number of the ectopic sites of the to-be-detected cucumber germplasm and a standard cucumber germplasm (to-be-detected cucumber germplasm) is more than 2, judging the to-be-detected cucumber germplasm and the standard cucumber germplasm as different cucumber germplasm; the greater the number of differential sites, the more distant the genetic relationship.

If the number of the different sites of the to-be-detected cucumber germplasm and a standard cucumber germplasm (to-be-detected cucumber germplasm) is 0-2, the to-be-detected cucumber germplasm and the standard cucumber germplasm are judged to be similar cucumber germplasm.

In a seventh aspect, the invention provides the above SNP sites, SNP primer combinations, SNP kits, DNA fingerprint database detection methods, for use in X1 or X2:

x1: identifying whether the germplasm of the cucumber to be detected belongs to one of standard cucumber germplasms;

x2: identifying the specific germplasm of the cucumber to be detected as the standard cucumber germplasm;

both X1 and X2 belong to the application of identifying the authenticity of cucumber germplasm, including identifying the authenticity of the cucumber germplasm.

The following examples are given to facilitate a better understanding of the invention, but do not limit the invention. The experimental procedures in the following examples are conventional unless otherwise specified. The test materials used in the following examples were purchased from a conventional biochemical reagent store unless otherwise specified.