阅读说明：本技术 一种基于Y-STR基因座和Y-indel基因座的复合扩增体系及其使用的引物组合 (Composite amplification system based on Y-STR locus and Y-indel locus and primer combination used by same ) 是由 尚蕾 丁光树 莫晓婷 余政梁 孙辉 吴俞衡 李万水 于 2019-11-19 设计创作，主要内容包括：本发明公开了一种基于Y-STR基因座和Y-indel基因座的复合扩增体系及其使用的引物组合。本发明所提供的引物组合由序列表中的序列1至序列74所示的74种DNA分子组成。采用所述引物组合构建复合扩增体系,然后进行男性个体的STR分型,检出的多态性高,在父系亲缘关系鉴定、家系排查等方面具有重要的鉴别价值。本发明具有重要的应用价值。(The invention discloses a composite amplification system based on a Y-STR locus and a Y-indel locus and a primer combination used by the same. The primer combination provided by the invention consists of 74 DNA molecules shown in a sequence 1 to a sequence 74 in a sequence table. The primer combination is adopted to construct a composite amplification system, then male STR typing is carried out, the detected polymorphism is high, and the method has important identification value in the aspects of paternal relationship identification, family investigation and the like. The invention has important application value.)

1. A primer set comprising primer 1, primer 2, primer 3, primer 4, primer 5, primer 6, primer 7, primer 8, primer 9, primer 10, primer 11, primer 12, primer 13, primer 14, primer 15, primer 16, primer 17, primer 18, primer 19, primer 20, primer 21, primer 22, primer 23, primer 24, primer 25, primer 26, primer 27, primer 28, primer 29, primer 30, primer 31, primer 32, primer 33, primer 34, primer 35, primer 36, primer 37, primer 38, primer 39, primer 40, primer 41, primer 42, primer 43, primer 44, primer 45, primer 46, primer 47, primer 48, primer 49, primer 50, primer 51, primer 52, primer 53, primer 54, primer 55, primer 56, primer 57, primer 58, primer 59, primer 60, primer 61, primer 62, primer 8, primer 9, primer 28, primer 13, primer 20, primer 16, primer 20, primer 25, primer 36, primer 36, primer 38, primer, primer 63, primer 64, primer 65, primer 66, primer 67, primer 68, primer 69, primer 70, primer 71, primer 72, primer 73 and primer 74;

primer 1 is SEQ ID NO: 1, a single-stranded DNA molecule;

primer 2 is SEQ ID NO: 2;

primer 3 is SEQ ID NO: 3, a single-stranded DNA molecule;

primer 4 is SEQ ID NO: 4, a single-stranded DNA molecule;

primer 5 is SEQ ID NO: 5, a single-stranded DNA molecule;

primer 6 is SEQ ID NO: 6;

primer 7 is SEQ ID NO: 7;

primer 8 is SEQ ID NO: 8, a single-stranded DNA molecule;

primer 9 is SEQ ID NO: 9, a single-stranded DNA molecule;

primer 10 is SEQ ID NO: 10, a single-stranded DNA molecule;

primer 11 is SEQ ID NO: 11;

primer 12 is SEQ ID NO: 12;

primer 13 is SEQ ID NO: 13, a single-stranded DNA molecule;

primer 14 is SEQ ID NO: 14, a single-stranded DNA molecule;

primer 15 is SEQ ID NO: 15, a single-stranded DNA molecule;

primer 16 is SEQ ID NO: 16;

primer 17 is SEQ ID NO: 17;

primer 18 is SEQ ID NO: 18, a single-stranded DNA molecule;

primer 19 is SEQ ID NO: 19;

primer 20 is SEQ ID NO: 20, a single-stranded DNA molecule;

primer 21 is SEQ ID NO: 21, a single-stranded DNA molecule;

primer 22 is SEQ ID NO: 22;

primer 23 is SEQ ID NO: 23;

primer 24 is SEQ ID NO: 24, a single-stranded DNA molecule;

primer 25 is SEQ ID NO: 25;

primer 26 is SEQ ID NO: 26;

primer 27 is SEQ ID NO: 27, a single-stranded DNA molecule;

primer 28 is SEQ ID NO: 28;

primer 29 is SEQ ID NO: 29;

primer 30 is SEQ ID NO: 30;

primer 31 is SEQ ID NO: 31;

primer 32 is SEQ ID NO: 32, a single-stranded DNA molecule;

primer 33 is SEQ ID NO: 33;

primer 34 is SEQ ID NO: 34, a single-stranded DNA molecule;

primer 35 is SEQ ID NO: 35;

primer 36 is SEQ ID NO: 36;

primer 37 is SEQ ID NO: 37;

primer 38 is SEQ ID NO: 38;

primer 39 is SEQ ID NO: 39;

primer 40 is SEQ ID NO: 40;

primer 41 is SEQ ID NO: 41;

primer 42 is SEQ ID NO: 42;

primer 43 is SEQ ID NO: 43;

primer 44 is SEQ ID NO: 44, a single-stranded DNA molecule;

primer 45 is SEQ ID NO: 45, a single-stranded DNA molecule;

primer 46 is SEQ ID NO: 46;

primer 47 is SEQ ID NO: 47;

primer 48 is SEQ ID NO: 48, a single-stranded DNA molecule;

primer 49 is SEQ ID NO: 49;

primer 50 is SEQ ID NO: 50;

primer 51 is SEQ ID NO: 51;

primer 52 is SEQ ID NO: 52;

primer 53 is SEQ ID NO: 53, a single-stranded DNA molecule;

primer 54 is SEQ ID NO: 54, or a single-stranded DNA molecule thereof;

primer 55 is SEQ ID NO: 55;

primer 56 is SEQ ID NO: 56;

primer 57 is SEQ ID NO: 57;

primer 58 is SEQ ID NO: 58;

primer 59 is SEQ ID NO: 59;

primer 60 is SEQ ID NO: 60;

primer 61 is SEQ ID NO: 61;

primer 62 is SEQ ID NO: 62;

primer 63 is SEQ ID NO: 63;

primer 64 is SEQ ID NO: 64;

primer 65 is SEQ ID NO: 65;

primer 66 is SEQ ID NO: 66;

primer 67 is SEQ ID NO: 67, a single stranded DNA molecule;

primer 68 is SEQ ID NO: 68;

primer 69 is SEQ ID NO: 69;

primer 70 is SEQ ID NO: 70;

primer 71 is SEQ ID NO: 71;

primer 72 is SEQ ID NO: 72;

primer 73 is SEQ ID NO: 73;

primer 74 is SEQ ID NO: 74, or a single-stranded DNA molecule as shown.

2. The primer combination of claim 1, wherein: primer 1, primer 2, primer 3, primer 4, primer 5, primer 6, primer 7, primer 8, primer 9, primer 10, primer 11, primer 12, primer 13, primer 14, primer 15, primer 16, primer 17, primer 18, primer 19, primer 20, primer 21, primer 22, primer 23, primer 24, primer 25, primer 26, primer 27, primer 28, primer 29, primer 30, primer 31, primer 32, primer 33, primer 34, primer 35, primer 36, primer 37, primer 38, primer 39, primer 40, primer 41, primer 42, primer 43, primer 44, primer 45, primer 46, primer 47, primer 48, primer 49, primer 50, primer 51, primer 52, primer 53, primer 54, primer 55, primer 56, primer 57, primer 58, primer 59, primer 60, primer 61, primer 62, primer 63, primer 64, primer 65, primer 52, primer 53, primer 54, primer 60, The molar ratio of primer 66, primer 67, primer 68, primer 69, primer 70, primer 71, primer 72, primer 73 and primer 74 was 150: 150: 70: 70: 140: 140: 180: 180: 170: 170: 300: 300: 160: 160: 130: 130: 130: 130: 120: 120: 80: 80: 90: 90: 140: 140: 180: 180: 100: 100: 150: 150: 180: 180: 210: 210: 120: 120: 200: 200: 210: 210: 160: 160: 400: 400: 180: 180: 120: 120: 120: 120: 200: 200: 200: 200: 250: 250: 400: 400: 150: 150: 230: 230: 130: 130: 190: 190: 210: 210: 140: 140: 300: 300.

3. the primer combination of claim 1 or 2, wherein: primer 1, primer 3, primer 5, primer 7, primer 9, primer 11, primer 13, primer 15, primer 17, primer 19, primer 21, primer 23, primer 25, primer 27, primer 29, primer 31, primer 33, primer 35, primer 37, primer 39, primer 41, primer 43, primer 45, primer 47, primer 49, primer 51, primer 53, primer 55, primer 57, primer 59, primer 61, primer 63, primer 65, primer 67, primer 69, primer 71 and primer 73 are all fluorescently labeled.

4. The primer combination of claim 3, wherein:

primer 1, primer 3, primer 5, primer 7, primer 9, primer 11, primer 13, primer 15 and primer 17 were labeled with FAM;

primer 19, primer 21, primer 23, primer 25, primer 27, primer 29 and primer 31 are labeled with HEX;

primer 33, primer 35, primer 37, primer 39, primer 41, primer 43 and primer 45 are labeled with TAMRA;

primer 47, primer 49, primer 51, primer 53, primer 55, primer 57 and primer 59 are labeled with ROX;

primer 61, primer 63, primer 65, primer 67, primer 69, primer 71 and primer 73 were labeled with SID.

5. A multiplex amplification system based on a Y-STR locus and a Y-Indels locus, comprising a primer combination according to any one of claims 1 to 4;

the Y-STR locus comprises DYS19, DYS385a/b, DYS389I/II, DYS390, DYS391, DYS392, DYS393, DYS437, DYS448, DYS456, DYS458, DYS635, Y _ GATA _ H4, DYS438, DYS439, DYS460, DYS533, DYS576, DYS449, DYS518, DYS 387S1, DYS627, DYS570, DYS447, DYS444, DYS549, DYS557, DYS643, DYS481, DYS596, DYS527a/b, DYS508, DYS593, and DYS 645;

the Y-Indels loci include rs771783753, rs199815934, and rs 759551978.

6. The multiplex amplification system of claim 5, wherein:

the concentration of the primer 1, the primer 2, the primer 31, the primer 32, the primer 61 and the primer 62 in the composite amplification system is 150 mM;

the concentration of the primer 3 and the primer 4 in the composite amplification system is 70 mM;

the concentration of the primer 5, the primer 6, the primer 25, the primer 26, the primer 71 and the primer 72 in the composite amplification system is 140 mM;

the concentration of primer 7, primer 8, primer 27, primer 28, primer 33, primer 34, primer 47 and primer 48 in the multiplex amplification system was 180 mM;

the concentration of the primer 9 and the primer 10 in the composite amplification system is 170 mM;

the concentration of the primer 11, the primer 12, the primer 73 and the primer 74 in the composite amplification system is 300 mM;

the concentration of primer 13, primer 14, primer 43 and primer 44 in the multiplex amplification system was 160 mM;

the concentration of the primer 15, the primer 16, the primer 17, the primer 18, the primer 65 and the primer 66 in the composite amplification system is 130 mM;

the concentration of primer 19, primer 20, primer 37, primer 38, primer 49, primer 50, primer 51 and primer 52 in the multiplex amplification system is 120 mM;

the concentration of the primer 21 and the primer 22 in the composite amplification system is 80 mM;

the concentration of the primer 23 and the primer 24 in the composite amplification system is 90 mM;

the concentration of the primer 29 and the primer 30 in the composite amplification system is 100 mM;

the concentration of the primer 35, the primer 36, the primer 41, the primer 42, the primer 69 and the primer 70 in the composite amplification system is 210 mM;

the concentration of the primer 39, the primer 40, the primer 53, the primer 54, the primer 55 and the primer 56 in the composite amplification system is 200 mM;

the concentration of the primer 45, the primer 46, the primer 59 and the primer 60 in the composite amplification system is 400 mM;

the concentration of primer 57 and primer 58 in the multiplex amplification system was 250 mM;

the concentration of the primer 63 and the primer 64 in the composite amplification system is 230 mM;

the concentration of primer 67 and primer 68 in the multiplex amplification system was 190 mM.

7. The multiplex amplification system of claim 5 or claim 6, wherein: the composite amplification system also comprises reagents required for carrying out PCR amplification reaction; the "reagents required for performing a PCR amplification reaction" does not include primers required for a PCR amplification reaction.

8. A kit comprising the primer combination of any one of claims 1 to 4; the kit is used for STR typing of male individuals.

9. A method for preparing the multiplex amplification system according to any one of claims 5 to 7 or the kit according to claim 8, comprising the step of packaging each primer in the primer combination according to any one of claims 1 to 4 separately.

10, X1) or X2):

x1) the use of the primer combination of any one of claims 1 to 4, or the multiplex amplification system of any one of claims 5 to 7, for the manufacture of a kit for STR typing of male individuals;

x2) the primer combination of any one of claims 1 to 4, or the multiplex amplification system of any one of claims 5 to 7, for use in STR typing of male individuals.

Technical Field

The invention belongs to the technical field of forensic medicine, and particularly relates to a composite amplification system based on Y-STR loci and Y-indel loci and a primer combination used by the same, in particular to a composite amplification system based on 38Y-STR loci and 3Y-indel loci and a primer combination used by the same.

Background

Short Tandem Repeat (STR) belongs to the second generation genetic marker, and has been widely applied to the fields of population genetics and forensic medicine due to the characteristics of small repeat unit, high mutation rate, wide distribution in genome and the like. In the human genome, there is one STR site approximately every 15-20kb, which accounts for 10% of the total human genome. These STRs usually consist of 2-6 bases in tandem repeat units, and due to the differences in repeat units and repeat times, they show great variability among people of different ethnic groups and regions, and are manifested as genetic polymorphisms.

The Y-STR genetic marker is an STR genetic marker distributed on human Y chromosome, is an effective means for testing and identifying in the court science field at present, and is an important supplement for autosomal STR, mitochondrial genetic markers and the like. Because the Y chromosome is peculiar to the male individual, most (more than 95 percent) of the Y chromosome is a non-recombinant region, the Y-STR presents the paternal inheritance characteristic and plays an important role in the aspects of paternal inheritance relationship identification, mixed-spot male individual inspection and the like. Meanwhile, by utilizing the characteristic that the Y-STR is relatively conserved in one family, the family of the important region can be subjected to Y-STR typing inspection to carry out family investigation, so that the investigation personnel are assisted to quickly lock the family of the suspect and the progress of the case is accelerated. In recent years, the Y-STR technology has helped to break down a large number of hit-case scenarios.

With the development of Y-STR technology, the amplification and inspection system related to Y-STR at home and abroad is continuously perfect, and the reagent products are also continuously abundant. Initially, the Y-STR amplification detection system can only detect a few Y-STR loci. Subsequently, the number of loci in the reagent System is increased continuously, and the detection efficiency is enhanced continuously, for example, the Promega company firstly provides a PowerPlex Y System containing 12Y-STR loci, and then develops and develops a PowerPlex Y23 System capable of detecting 23Y-STR loci; the ABI company provides a Yfiler kit containing 17Y-STR loci, and then develops Yfiler Plus capable of detecting 27Y-STR loci; in China, DNATYPERY series reagents are independently developed by the material evidence identification center of the Ministry of public Security, and Y-STR detection reagents are continuously proposed by Texas China Union biotechnology Limited and Suzhou Ministry of Japan and GenBank technology Limited, so that six-color fluorescent amplification detection products containing more than 30Y-STR loci are available at present. The product can play a certain identification role in the inspection materials for male and female component mixing or multiple male component mixing, and can assist in searching the family of the criminal suspect, thereby greatly reducing the investigation range. However, existing kit products show deficiencies in both locus number and locus mutation rate, and thus in some case applications, male individuals cannot be effectively distinguished. Based on this, the Y-STR test reagent product should contain more loci, and especially should contain 20Y-STR core loci (such as DYS19, DYS385a/b, DYS389I/II, DYS390, DYS391, DYS392, DYS393, DYS437, DYS448, DYS456, DYS458, DYS635, Y GATA H4, DYS438, DYS439, DYS460, DYS533, and DYS576) selected by the ministry for the construction of the Y database and 15 preferred loci (such as DYS449, DYS518, DYS 387S1, DYS627, DYS570, DYS447, DYS444, DYS557, DYS481, DYS645, DYS527a/b) selected by the ministry of Shandong) for the construction of the Y database, so as to achieve a greater efficacy of the DYS593 and DYS549 in the practical test system.

Disclosure of Invention

The invention aims to perform STR typing on male individuals.

The primer combination is protected firstly, and can comprise primer 1, primer 2, primer 3, primer 4, primer 5, primer 6, primer 7, primer 8, primer 9, primer 10, primer 11, primer 12, primer 13, primer 14, primer 15, primer 16, primer 17, primer 18, primer 19, primer 20, primer 21, primer 22, primer 23, primer 24, primer 25, primer 26, primer 27, primer 28, primer 29, primer 30, primer 31, primer 32, primer 33, primer 34, primer 35, primer 36, primer 37, primer 38, primer 39, primer 40, primer 41, primer 42, primer 43, primer 44, primer 45, primer 46, primer 47, primer 48, primer 49, primer 50, primer 51, primer 52, primer 53, primer 54, primer 55, primer 56, primer 57, primer 58, primer 59, primer 60, primer 61, primer 55, primer 56, primer 59, primer 60, Primer 62, primer 63, primer 64, primer 65, primer 66, primer 67, primer 68, primer 69, primer 70, primer 71, primer 72, primer 73 and primer 74;

primer 1 can be SEQ ID NO: 1, a single-stranded DNA molecule;

primer 2 can be SEQ ID NO: 2;

primer 3 can be SEQ ID NO: 3, a single-stranded DNA molecule;

primer 4 can be SEQ ID NO: 4, a single-stranded DNA molecule;

primer 5 can be SEQ ID NO: 5, a single-stranded DNA molecule;

primer 6 can be SEQ ID NO: 6;

primer 7 can be SEQ ID NO: 7;

primer 8 can be SEQ ID NO: 8, a single-stranded DNA molecule;

primer 9 can be SEQ ID NO: 9, a single-stranded DNA molecule;

primer 10 can be SEQ ID NO: 10, a single-stranded DNA molecule;

primer 11 can be SEQ ID NO: 11;

primer 12 can be SEQ ID NO: 12;

primer 13 can be SEQ ID NO: 13, a single-stranded DNA molecule;

primer 14 can be SEQ ID NO: 14, a single-stranded DNA molecule;

primer 15 can be SEQ ID NO: 15, a single-stranded DNA molecule;

primer 16 can be SEQ ID NO: 16;

primer 17 can be SEQ ID NO: 17;

primer 18 can be SEQ ID NO: 18, a single-stranded DNA molecule;

primer 19 can be SEQ ID NO: 19;

primer 20 can be SEQ ID NO: 20, a single-stranded DNA molecule;

primer 21 can be SEQ ID NO: 21, a single-stranded DNA molecule;

primer 22 can be SEQ ID NO: 22;

primer 23 can be SEQ ID NO: 23;

primer 24 can be SEQ ID NO: 24, a single-stranded DNA molecule;

primer 25 can be SEQ ID NO: 25;

primer 26 can be SEQ ID NO: 26;

primer 27 can be SEQ ID NO: 27, a single-stranded DNA molecule;

primer 28 may be SEQ ID NO: 28;

primer 29 can be SEQ ID NO: 29;

the primer 30 may be SEQ ID NO: 30;

primer 31 can be SEQ ID NO: 31;

primer 32 can be SEQ ID NO: 32, a single-stranded DNA molecule;

primer 33 may be SEQ ID NO: 33;

primer 34 can be SEQ ID NO: 34, a single-stranded DNA molecule;

primer 35 can be SEQ ID NO: 35;

primer 36 can be SEQ ID NO: 36;

primer 37 may be SEQ ID NO: 37;

primer 38 can be SEQ ID NO: 38;

primer 39 can be SEQ ID NO: 39;

primer 40 can be SEQ ID NO: 40;

primer 41 can be SEQ ID NO: 41;

primer 42 can be SEQ ID NO: 42;

primer 43 can be SEQ ID NO: 43;

primer 44 may be SEQ ID NO: 44, a single-stranded DNA molecule;

primer 45 can be SEQ ID NO: 45, a single-stranded DNA molecule;

primer 46 may be SEQ ID NO: 46;

primer 47 can be SEQ ID NO: 47;

primer 48 can be SEQ ID NO: 48, a single-stranded DNA molecule;

primer 49 can be SEQ ID NO: 49;

primer 50 can be SEQ ID NO: 50;

primer 51 may be SEQ ID NO: 51;

primer 52 can be SEQ ID NO: 52;

primer 53 may be SEQ ID NO: 53, a single-stranded DNA molecule;

primer 54 can be SEQ ID NO: 54, or a single-stranded DNA molecule thereof;

primer 55 can be SEQ ID NO: 55;

primer 56 can be SEQ ID NO: 56;

primer 57 can be SEQ ID NO: 57;

primer 58 can be SEQ ID NO: 58;

primer 59 can be SEQ ID NO: 59;

primer 60 can be SEQ ID NO: 60;

primer 61 can be SEQ ID NO: 61;

primer 62 can be SEQ ID NO: 62;

primer 63 may be SEQ ID NO: 63;

primer 64 can be SEQ ID NO: 64;

primer 65 may be SEQ ID NO: 65;

primer 66 can be SEQ ID NO: 66;

primer 67 may be SEQ ID NO: 67, a single stranded DNA molecule;

primer 68 can be SEQ ID NO: 68;

primer 69 can be SEQ ID NO: 69;

primer 70 can be SEQ ID NO: 70;

primer 71 may be SEQ ID NO: 71;

primer 72 can be SEQ ID NO: 72;

primer 73 can be SEQ ID NO: 73;

primer 74 can be SEQ ID NO: 74, or a single-stranded DNA molecule as shown.

The primer combination may specifically include primer 1, primer 2, primer 3, primer 4, primer 5, primer 6, primer 7, primer 8, primer 9, primer 10, primer 11, primer 12, primer 13, primer 14, primer 15, primer 16, primer 17, primer 18, primer 19, primer 20, primer 21, primer 22, primer 23, primer 24, primer 25, primer 26, primer 27, primer 28, primer 29, primer 30, primer 31, primer 32, primer 33, primer 34, primer 35, primer 36, primer 37, primer 38, primer 39, primer 40, primer 41, primer 42, primer 43, primer 44, primer 45, primer 46, primer 47, primer 48, primer 49, primer 50, primer 51, primer 52, primer 53, primer 54, primer 55, primer 56, primer 57, primer 58, primer 59, primer 60, primer 61, primer 62, primer 8, primer 9, primer 28, primer 25, primer 13, primer 35, primer 25, primer 60, primer 59, primer, primer 63, primer 64, primer 65, primer 66, primer 67, primer 68, primer 69, primer 70, primer 71, primer 72, primer 73 and primer 74.

In any of the above-mentioned primer combinations, primer 1, primer 2, primer 3, primer 4, primer 5, primer 6, primer 7, primer 8, primer 9, primer 10, primer 11, primer 12, primer 13, primer 14, primer 15, primer 16, primer 17, primer 18, primer 19, primer 20, primer 21, primer 22, primer 23, primer 24, primer 25, primer 26, primer 27, primer 28, primer 29, primer 30, primer 31, primer 32, primer 33, primer 34, primer 35, primer 36, primer 37, primer 38, primer 39, primer 40, primer 41, primer 42, primer 43, primer 44, primer 45, primer 46, primer 47, primer 48, primer 49, primer 50, primer 51, primer 52, primer 53, primer 54, primer 55, primer 56, primer 57, primer 58, primer 59, primer 60, primer 61, primer 56, primer 7, primer 25, primer 27, primer 28, primer 25, primer 50, primer 52, primer 54, primer 55, primer 59, primer, The molar ratio of primer 62, primer 63, primer 64, primer 65, primer 66, primer 67, primer 68, primer 69, primer 70, primer 71, primer 72, primer 73, and primer 74 can be 150: 150: 70: 70: 140: 140: 180: 180: 170: 170: 300: 300: 160: 160: 130: 130: 130: 130: 120: 120: 80: 80: 90: 90: 140: 140: 180: 180: 100: 100: 150: 150: 180: 180: 210: 210: 120: 120: 200: 200: 210: 210: 160: 160: 400: 400: 180: 180: 120: 120: 120: 120: 200: 200: 200: 200: 250: 250: 400: 400: 150: 150: 230: 230: 130: 130: 190: 190: 210: 210: 140: 140: 300: 300.

in any of the primer combinations described above, primer 1, primer 3, primer 5, primer 7, primer 9, primer 11, primer 13, primer 15, primer 17, primer 19, primer 21, primer 23, primer 25, primer 27, primer 29, primer 31, primer 33, primer 35, primer 37, primer 39, primer 41, primer 43, primer 45, primer 47, primer 49, primer 51, primer 53, primer 55, primer 57, primer 59, primer 61, primer 63, primer 65, primer 67, primer 69, primer 71 and primer 73 may be labeled with a fluorescent label.

In any of the primer combinations described above, primer 1, primer 3, primer 5, primer 7, primer 9, primer 11, primer 13, primer 15 and primer 17 may be labeled with FAM. Primer 19, primer 21, primer 23, primer 25, primer 27, primer 29 and primer 31 may be labeled with HEX. Primer 33, primer 35, primer 37, primer 39, primer 41, primer 43 and primer 45 may be labeled with TAMRA. Primer 47, primer 49, primer 51, primer 53, primer 55, primer 57 and primer 59 may be labeled with ROX. Primer 61, primer 63, primer 65, primer 67, primer 69, primer 71 and primer 73 can be labeled with SID. The 5' ends of primer 1, primer 3, primer 5, primer 7, primer 9, primer 11, primer 13, primer 15 and primer 17 may be labeled with FAM. The 5' ends of primer 19, primer 21, primer 23, primer 25, primer 27, primer 29 and primer 31 may be labeled with HEX. The 5' ends of primer 33, primer 35, primer 37, primer 39, primer 41, primer 43 and primer 45 may be labeled with TAMRA. The 5' ends of primer 47, primer 49, primer 51, primer 53, primer 55, primer 57 and primer 59 may be labeled with ROX. The 5' ends of primer 61, primer 63, primer 65, primer 67, primer 69, primer 71 and primer 73 may be labeled with SID.

The invention also discloses a composite amplification system based on the Y-STR locus and the Y-Indels locus, which can comprise any one of the primer combinations;

the Y-STR locus may comprise DYS19, DYS385a/b, DYS389I/II, DYS390, DYS391, DYS392, DYS393, DYS437, DYS448, DYS456, DYS458, DYS635, Y _ GATA _ H4, DYS438, DYS439, DYS460, DYS533, DYS576, DYS449, DYS518, DYS 387S1, DYS627, DYS570, DYS447, DYS444, DYS549, DYS557, DYS643, DYS481, DYS596, DYS527a/b, DYS508, DYS593, and DYS 645;

the Y-Indels locus may comprise rs771783753, rs199815934 and rs 759551978.

The Y-STR locus may specifically consist of DYS19, DYS385a/b, DYS389I/II, DYS390, DYS391, DYS392, DYS393, DYS437, DYS448, DYS456, DYS458, DYS635, Y _ GATA _ H4, DYS438, DYS439, DYS460, DYS533, DYS576, DYS449, DYS518, DYS 387S1, DYS627, DYS570, DYS447, DYS444, DYS549, DYS557, DYS643, DYS481, DYS596, DYS527a/b, DYS508, DYS593, and DYS.

The Y-Indels locus may specifically consist of rs771783753, rs199815934 and rs 759551978.

DYS576, DYS449, DYS518, DYF387S1, DYS627 and DYS570 are rapid mutating Y-STR loci.

The 41 loci (namely 38Y-STR loci and 3Y-Indels loci) comprise 20Y-STR core loci and 15 preferred loci selected by the Ministry of public security for Y database construction, and also comprise DYS593 and DYS645 selected by the Ministry of public security of Shandong for Y database construction.

The composite amplification system can specifically consist of any one of the primer combinations.

In any of the above multiplex amplification systems, the concentration of primer 1, primer 2, primer 31, primer 32, primer 61 and primer 62 in the multiplex amplification system can be 150 mM. The concentration of each of primer 3 and primer 4 in the multiplex amplification system may be 70 mM. Primer 5, primer 6, primer 25, primer 26, primer 71 and primer 72 may each be present in the multiplex amplification system at a concentration of 140 mM. The concentration of primer 7, primer 8, primer 27, primer 28, primer 33, primer 34, primer 47 and primer 48 in the multiplex amplification system may be 180mM each. The concentration of each of primer 9 and primer 10 in the multiplex amplification system may be 170 mM. The concentration of primer 11, primer 12, primer 73 and primer 74 in the multiplex amplification system may be 300mM each. The concentration of each of primer 13, primer 14, primer 43 and primer 44 in the multiplex amplification system may be 160 mM. Primer 15, primer 16, primer 17, primer 18, primer 65 and primer 66 may each be present in the multiplex amplification system at a concentration of 130 mM. The concentration of each of primer 19, primer 20, primer 37, primer 38, primer 49, primer 50, primer 51 and primer 52 in the multiplex amplification system may be 120 mM. The concentration of each of primer 21 and primer 22 in the multiplex amplification system may be 80 mM. The concentration of each of the primer 23 and the primer 24 in the multiplex amplification system may be 90 mM. The concentration of each of the primer 29 and the primer 30 in the multiplex amplification system may be 100 mM. The concentration of primer 35, primer 36, primer 41, primer 42, primer 69 and primer 70 in the multiplex amplification system may each be 210 mM. The concentration of each of primer 39, primer 40, primer 53, primer 54, primer 55 and primer 56 in the multiplex amplification system may be 200 mM. The concentration of primer 45, primer 46, primer 59, and primer 60 in the multiplex amplification system may each be 400 mM. The concentration of each of the primer 57 and the primer 58 in the multiplex amplification system may be 250 mM. The concentration of each of the primers 63 and 64 in the multiplex amplification system may be 230 mM. The concentration of each of primer 67 and primer 68 in the multiplex amplification system may be 190 mM.

In the above composite amplification system, the composite amplification system may further comprise reagents required for performing a PCR amplification reaction; the "reagents required for performing a PCR amplification reaction" does not include primers required for a PCR amplification reaction.

The "reagents required for carrying out PCR amplification reaction" may include DNA polymerase, dNTP, Mg ²⁺At least one of BSA, KCl and Tris.

The concentration of the DNA polymerase in the multiplex amplification system may be 0.1U/. mu.L. The concentration of the dNTPs in the multiplex amplification system can be 200. mu.M (i.e., the concentration of dATP, dTTP, dCTP, and dGTP is 200. mu.M). The Mg ²⁺The concentration in the multiplex amplification system may be 1.5 mM. The concentration of the BSA in the multiplex amplification system may be 0.8 mg/mL. The concentration of the KCl in the multiplex amplification system can be 50 mM. The concentration of the Tris in the multiplex amplification system may be 10 mM.

The composite amplification system can specifically comprise any one of the primer combinations and reagents required for carrying out PCR amplification reaction.

A kit containing any of the primer combinations also belongs to the protection scope of the invention. The kit is used for STR typing of male individuals.

The preparation method of any one of the above composite amplification systems or the kit containing any one of the above primer combinations also belongs to the protection scope of the invention.

The method for preparing the multiplex amplification system or the kit containing the primer combination may comprise the step of packaging each primer in any one of the primer combinations separately.

The following X1) or X2) also belong to the scope of the present invention.

X1) any one of the primer combinations or the multiplex amplification system, and the application thereof in preparing a kit for STR typing of male individuals.

X2) any one of the primer combinations or the composite amplification system, and the application thereof in STR typing of male individuals.

Any one of the STR typing may be Y-STR typing.

Any of the above male individuals may be chinese male individuals.

The composite amplification system provided by the invention is adopted to carry out STR typing on a sample I (9948 human genome DNA standard) or a sample II (human (known as male) blood collection card). The result shows that the first sample and the second sample both obtain complete STR typing, and the peaks are sharp and clear, the balance is good, and no Pull-up peak, stutter band and non-specific amplification product appear. 9948 the results of the human genome DNA standard also show that the typing results of the composite amplification system for 38Y-STR loci and 3Y-Indels loci shown in Table 1 are correct, and the requirements of forensic Y-STR inspection can be completely met. Therefore, the composite amplification system provided by the invention can be used for amplifying 38Y-STR loci (7 of which are rapid mutation Y-STR loci) and 3Y-Indels loci, and the composite amplification system can obtain 41 different allele fragments at most by one-time amplification and has very high amplification efficiency. Meanwhile, the composite amplification system contains 7 rapidly mutated Y-STR loci, and the detection efficiency is very strong. The composite amplification system provided by the invention has important application value in the aspects of paternal genetic relationship identification, family investigation and the like.

Drawings

FIG. 1 is a DNA detection map of sample one.

FIG. 2 is a DNA detection map of sample two.

Detailed Description

The present invention is described in further detail below with reference to specific embodiments, which are given for the purpose of illustration only and are not intended to limit the scope of the invention.

The experimental procedures in the following examples are conventional unless otherwise specified.

Materials, reagents and the like used in the following examples are commercially available unless otherwise specified.