阅读说明：本技术 一种鉴定小麦优势变异染色体及其遗传效应的方法 (Method for identifying wheat dominant variation chromosome and genetic effect thereof ) 是由 亓增军 吴楠 刘鑫 杨阳 王从磊 何梓铭 方佳欣 于 2021-08-06 设计创作，主要内容包括：本发明公开了一种鉴定小麦优势变异染色体及其遗传效应的方法,综合寡核苷酸探针荧光原位杂交、小麦芯片分析、混合分离群体表型比较和BSR-seq分析揭示小麦变异染色体遗传传递特点以及对基因组分化、基因表达和表型的效应,为发掘和利用小麦优势染色体变异及其关键基因(簇)进行小麦品种改良提供了新方案,属于作物遗传育种与农业生物技术领域。(The invention discloses a method for identifying a wheat dominant variant chromosome and a genetic effect thereof, which integrates oligonucleotide probe fluorescence in-situ hybridization, wheat chip analysis, mixed segregation population phenotype comparison and BSR-seq analysis to reveal genetic transmission characteristics of the wheat variant chromosome and the effects on genome differentiation, gene expression and phenotype, provides a new scheme for exploring and utilizing the wheat dominant chromosome variant and key genes (clusters) thereof to improve wheat varieties, and belongs to the technical field of crop genetic breeding and agricultural biology.)

1. A method for identifying wheat dominant variant chromosome and genetic effect thereof comprises the following steps:

(1) the method is characterized in that an oligonucleotide probe set ONMP #4 is used for identifying variant chromosomes (including structural variant chromosomes and polymorphic chromosomes) in Yangmai 158(Y158) and Fangshan wheat (FSXM), and five pairs of chromosomes are found to have differences, namely: 4A-3 vs 4A-1, 5A-4 vs 5A-1, 4B-2 vs 4B-1, perInv6B vs 6B-1, 7B-1 vs 7B-7;

(2) for the constructed Yangmai 158/mountain-turning wheat hybrid F₂Population, Recombinant Inbred Line (RIL) F₈Generation group and RIL-F₈The chromosome composition analysis of the inbred population of the heterozygous individual revealed the genetic transmission of the variant chromosomes identified in (1), and the chi-square test was performed, except that perInv6B was partially separated in both populations and 4A-3 was performed at F₂In addition to segregation in the population, the remaining variant chromosomes are normally segregated, but in RIL-F₈The heterozygous individual plants are bred into a progeny group, and the PerInv6B is normally separated;

(3) the Y158/FSXM RIL-F in (2)₈Carrying out genotype analysis on a wheat 15K chip, and carrying out genetic differentiation index analysis (Fst) on five pairs of clusters divided according to the types of wheat variant chromosomes to find that difference SNP (single nucleotide polymorphism) intervals exist between two pairs of clusters of PerInv6B vs 6B-1 and 5A-4 vs 5A-1 (figure 4) at the corresponding inversed positions or near polymorphic segments, wherein the difference segments are 367.05Mb and 51.97Mb respectively;

(4) for Y158/FSXM RIL-F₁₀Dividing the chromosome into two mixed separation groups based on chromosome variation types, and finding that 21 genes are differentially expressed between parent and PerInv6B and 6B-1 mixed pools and relate to genes such as alpha-amylase, serine, threonine protein kinase, peroxidase and the like;

(5) using Y158/FSXM RIL-F₁₀Two-point and three-repetition phenotypic data analysis shows that PerInv6B, 5A-4 and 3B-2 respectively have the effects of reducing plant height, increasing grain number per spike, increasing grain length per spike and the like under specific environments.

2. Use of the method of claim 1 in the identification of dominant variant chromosomes in wheat and key genes (clusters) thereof.

3. Use of the method of claim 1 in wheat breeding.

One, the technical field

The patent discloses a method for identifying dominant variant chromosomes and genetic effects of the dominant variant chromosomes of wheat, and belongs to the technical field of crop genetic breeding and agricultural biology.

Second, background Art

Common wheat is a typical allohexaploid species, whose Genome is significantly mutated in order to increase adaptability to stress during polyploidization, domestication and modification (see references: Badaeva E, Dedkova O, Gay G, Pukhalskyi V, Zelenin A, Bernard S, Bernard M. chromosoma reaangement in wheat: the hair types and distribution [ J ]. Genome, 2007, 50: 907-. With the rapid development of crop genomics, great progress was made in the analysis of genomic variation characteristics during wheat domestication and manual selection at the Molecular level, identifying a set of selectable marker sites and segments (see references: Hao C, Wang Y, Chao S, Li T, Liu H, Wang L, Zhang X. the iSelect 9K SNP analyzed transformed polyploidy mutation and intense human selection using transformed chromosome in wheat straw [ J ]. Scientific Reports, 2017, 7: 41; Hao C, Jiano C, Hou J, Li T, Liu H, Wang Y, Zhang J, Liu H, Bi Z, Xu F, Zhao J, Ma L, Y, joined U, Liyang X, applied R, sample R, Molecular analysis [ 1J ],247, Molecular analysis, 2020, 13: 1-19), which provides important reference for genome breeding based on omics, but the formation mechanism of the selectable marker and the segment thereof is still lack of deep knowledge, for example, the relationship between the chromosome variation and the evolution of wheat genome, and the specific effects of the chromosome variation on agronomic traits, adversity stress and gene expression regulation are only rarely and directly reported, wherein the great reason is that the current omics technology is difficult to sequence and assemble heterochromatin segments rich in highly repetitive sequences, so the identification of large-scale chromosome variation, especially reciprocal translocation, inversion, recessive aneuploidy and the like, is still dependent on cytogenetic analysis, but the early lack of simple and efficient chromosome identification technology makes the large-scale analysis of chromosome variation in the evolution of wheat varieties also have great difficulty; secondly, a technical system for integrating chromosome variation, plant genome evolution and genetic effect analysis into a system research is lacked, so that the current omics research and cytogenetic research are not combined with each other in many aspects.

Chromosomal variations reduce gene flow by either reducing fertility of the hybrids or by inhibiting recombination, thus causing reproductive segregation and the formation of new species (see reference: Schubert I. chromosome evolution [ J.)].Current opinion in plant biology，2007，10：109-115；Murat F，Xu J，Tannier E，Abrouk M，Guilhot N，Pont C，Messing J，Salse J.Ancestral grass karyotype reconstruction unravels new mechanisms of genome shuffling as a source of plant evolution[J]Genome research, 2010, 20: 1545-1557). The structural variation of chromosome has important influence on evolution and human disease occurrence due to the rearrangement of large-fragment DNA sequence, and is studied more deeply in animals, microorganisms and humans (de Joger R, Bolton M, Kombrink A, van den Berg G, Yadeta K, Thomma B. extended chromosome restriction driving evolution of viral in an anaerobic disease [ J.].Genome research，2013，23：1271-1282；Collins R，Brand H，Karczewski K，Zhao X，J，Francioli L，Khera A，Lowther C，Gauthier L，Wang H，Watts N，Solomonson M，O’Donnell A，Baumann A，Munshi R，Walker M，Whelan C，Huang Y，Brookings T，Sharpe T，Stone M，Valkanas E，Fu J，Tiao G，Laricchia K，Ruano V，Stevens C，Gupta N，Cusick C，Margolin L，Genome Aggregation Database Production T，Genome Aggregation Database C，Taylor K，Lin H，Rich S，Post W，Chen Y，Rotter J，Nusbaum C，Philippakis A，Lander E，Gabriel S，Neale B，Kathiresan S，Daly M，Banks E，MacArthur D，Talkowski M.A structural variation reference for medical and population genetics[J]Nature, 2020, 581: 444-451). However, the research on the plant is less, and particularly, the plant is more suitable for crops with huge genome, such as wheatThere is a lack of intensive research. The combination of genomics and cytogenetics promotes the research of chromosome variation mechanism and effect, and particularly, oligonucleotide probes developed based on genome sequencing provide powerful tools for clearly identifying large-range chromosome structure variation, and greatly promote the research progress of wheat chromosome structure and function. In this way, 8 oligonucleotide probe sets were successfully developed by the former research of the Nanjing university of agriculture for wheat (see references: Du P, Zhuang L, Wang Y, Yuan L, Wang Q, Wang D, Dawadondup, Tan L, Shen J, Xu H, ZHao H, Chu C, Qi Z. development of oligonucleotide and multiplex probes for nucleic and acid identification of wheat and tomato beta-amyloid [ J ] P]Genome, 2017, 60: 93-103), barley (see reference: zhang S, Zhu M, Shang Y, Wang J, Dawadundup, Zhuang L, Zhang J, Chu C, Qi z. physical organization of redundant sequences and chromosome diversity of barrel modified by Fluorescence In Situ Hybridization (FISH) [ J]Genome, 2019, 1: 329), maize (see references: zhu M, Du P, Zhuang L, Chu C, Qi Z.A simple and effective non-condensing FISH method for main chromosome diffusion using single-strand and oligonucleotide probes [ J]Genome, 2017, 60 (8): 657-: wu N, Li M, Sun H, Cao Z, Liu P, Ding T, Xu H, Chu C, Zhuang L, Qi Z.RNA-seq surfactants definition of chromosome-specific markers and transfer of eye chromosome to heat [ J]Molecular Breeding, 2018, 38: 6) thinopyrum bessarabicum (see reference: chen J, Tang Y, Yao L, Wu H, Tu X, Zhuang L, Qi Z. cytologic and molecular characterization of Thinopyrum bessaracinum chromogenes and structural rearrargements endogenous in steamed [ J]Molecular Breeding, 2019, 39: 146) and the chromosome identification of species, wherein the oligonucleotide probe set #4(ONPM #4) developed in wheat is utilized in the chromosome identification of wheat and related species thereof in a large amount due to simplicity and high efficiency, and becomes an important tool for identifying the chromosome variation of wheat (see reference: huang X, Zhu M, Zhuang L, Zhuang S, Wang J, Chen X, Wang D，Chen J，Bao Y，Guo J，Zhang J，Feng Y，Chu C，Du P，Qi Z，Wang H，Chen P.Structural chromosome rearrangements and polymorphisms identified in Chinese wheat cultivars by high-resolution multiplex oligonucleotide FISH[J]The therapeutic and Applied Genetics, 2018, 131: 1967-; wheat 'Chinese spring' aneuploid high-definition karyotype and use [ J ] based on oligonucleotide probe-sleeving padding]Crop academic newspaper, 2017, 43: 1575) and 1587) identifying a large number of structural variation and polymorphism chromosomes from Chinese wheat bred varieties and local varieties, wherein part of the variation chromosomes are derived from backbone parents and are transmitted frequently in derived varieties, and simultaneously identifying a large number of high-frequency polymorphism types which possibly represent dominant variation types of Chinese wheat, but an effective method is not available for identifying the genetic effect of the variation chromosomes, so that the dominant chromosome variation contained in wheat germplasm is difficult to determine and discover, and the application in breeding is limited.

Aiming at the problems, the invention provides a method for accurately identifying wheat mutation chromosomes by using oligonucleotide probe fluorescence in-situ hybridization, constructing a mixed separation population based on different mutation chromosome types by using a genetic population, identifying wheat dominant mutation chromosomes and revealing the effects of the mutation chromosomes on genome differentiation, phenotypic characters and gene expression regulation and control by using wheat chip hybridization, phenotypic identification and mixed pool transcriptome RNA-seq (BSR-seq) analysis, and providing a new scheme for researching and utilizing the wheat dominant mutation chromosomes and key genes (clusters) thereof to improve and evolve wheat varieties.

Third, the invention

The technical problem is as follows:

the invention aims to disclose a method for identifying dominant variant chromosomes and genetic effects of the dominant variant chromosomes.

The technical scheme is as follows:

1. in order to determine the type of wheat variant chromosome, oligonucleotide probe set FISH analysis is carried out on parents A and B based on the variant chromosome segregation population, and parents are compared to haveThe polymorphic or structural variation types of the differences are summarized as the signal distribution characteristics of the different segments of the paired variant chromosomes. Oligonucleotide probes using probe set ONPM # 4: repetitive sequences pSc119.2-1 and (GAA) comprising a 6-carboxyfluorescein (6-carboxyfluorescein, FAM, green) modification₁₀And 6-carboxytetramethylrhodamine (6-carboxytetramethylrhodamine, TAMRA, Red) modified repeat sequences pAs1-1, pAs1-3, pAs1-4, pAs1-6, AFA-3 and AFA-4. All probe sequences are referenced to Du P, Zhuang L, Wang Y, Yuan L, Wang Q, Wang D, Dawadondup, Tan L, Shen J, Xu H, ZHao H, Chu C, Qi Z.development of oligonucleotides and multiplex probes for probes and primers identification of wheat and thinopum Bessarabacter chromogenes [ J.P. ]].Genome，2017，60：93-103。

2. In order to reveal the genetic transmission characteristics of wheat variant chromosome, firstly, a hybrid F of parent A/parent B related to the variant chromosome is constructed₂To F₂Carrying out ONMP # 4-based oligonucleotide probe set FISH analysis on the single plant to obtain a hybrid F of the variant chromosome₂The distribution in the method is identified, the homozygous number and the heterozygous number of various variation types are analyzed, chi-square test is carried out on the variation types, and the genetic transmission characteristics are further analyzed. Secondly, chromosome composition analysis is carried out on the families in the high-generation line population of the Recombined Inbred Lines (RIL) with the same combination, and variant chromosomes related to different families are determined. The identified families were divided into different pairwise groups according to the type of chromosomal variation.

3. To investigate the population differentiation characteristics between different variant chromosome populations, the above identified population of parental A/parental B RIL high generation lines was genotyped in a wheat breeding chip, and the paired populations divided according to the wheat variant chromosome types were subjected to genetic differentiation index Analysis (Fst) (see references: Danecek P, Auton A, Absesas G, Albers C, Bank E, DePrist M, Handsaker R, Lunter G, Marth G, Sherry S, McVean G, Durbin R, genome Pronalysis G. the variant Analysis G. the variant call format and VCFtools [ J ]. Bioinformatics, 27: 2156-, 2020, 11: 5085) it is found that there are different SNP regions in the vicinity of the corresponding structural variation or polymorphism region between the paired clusters, and the size of the different region is determined. Genes within the interval were analyzed for their function in relation to molecular function, cellular composition, biological processes, etc. (see references: Tian T, Liu Y, Yan H, You Q, Yi X, Du Z, Xu W, Su Z. agriGO v 2.0: a GO analysis toolkit for the aggregative community, 2017 update [ J ]. Nucleic Acids Research, 2017, 45: 122-.

4. To analyze the effect of variant chromosomes on gene expression, BSR-Seq analysis was performed on the parental A/parental B RIL high generation population based on the type of chromosomal variation (see references: Wang Y, Xie J, Zhang H, Guo B, Ning S, Chen Y, Lu P, Wu Q, Li M, Zhang D, Guo G, Zhang Y, Liu D, Zou S, Tang J, Zhao H, Wang X, Li J, Yang W, Cao T, Yin G, Liu Z. mapping strain concrete yield gene YrZH22 in Chinee wot gene fusion Zhou ai 22 by bulk segregant RNA-Seq (BSR-Seq) and compatible genes analysis [ J. Applied, 130: 2011, SNP-related region-SNP-map-related region-SNP-related region-1, demarest B, Bisgrove B, Gorsi B, Su Y-C, Yost h. mmappr: tissue mapping analysis pipeline for porous RNA-seq [ J ]. Genome research, 2013, 23: 687-; ramirez R, Segovia V, Bird N, Fenwick P, Holdgate S, Berry S, Jack P, Camcamo M, Uauy C.RNA-Seq bulked segregant analysis enables the identification of high-resolution genetic markers for weaving in hexagonal leather [ J ]. Plant Biotechnology Journal, 2014, 13: 613-: FDR < 0.05, fold difference greater than 2 fold), which was annotated for function.

5. In order to reveal the effect of the variant chromosome on the wheat phenotype, a parent A/parent B RIL high generation line population is used for carrying out multi-year multi-point phenotype data investigation, the phenotype data of the family is divided into paired groups based on the chromosome variant type, and the IBM SPSS staticisics 22 is used for carrying out T-test analysis to clarify the influence of the variant chromosome on the phenotype.

Has the advantages that:

1. the ONMP # 4-based fluorescence in situ hybridization disclosed by the patent can clearly reveal wheat variant chromosomes, and provides important reference for sequencing and sequence assembly of centromere, telomere and other heterochromatin regions rich in highly-concatenated repetitive sequences;

2. the method for revealing the SNP difference interval between the variant chromosome groups by utilizing the wheat chip technology can quickly identify the differentiation of the variant chromosomes from the molecular level, is combined with the cytological result to assist in judging the break points of the variation, identifies the functional genes and metabolic pathways in the difference interval, and provides convenience for the research on the genome differentiation, the variety origin and evolution and the discovery of selective gene segments caused by the variation of the repetitive sequence;

3. the method for identifying the differential genes and phenotypes based on the mixed pool transcriptome sequencing of the wheat variant chromosomes provides a new scheme for discovering advantageous genes (clusters) related to dominant variant chromosomes and key sections thereof, and provides important information for the research of wheat origin and evolution, the variety improvement and the cultivation of backbone parents;

4. the method for identifying the dominant variant chromosome and the genetic effect of the dominant variant chromosome can be used for wheat natural populations and other plants, and provides an important reference for discovering the dominant variant chromosome and key trait genes of the plants.

Description of the drawings

FIG. 1: identification of Yangmai 158(Y158) and Fangshan wheat (FSXM) karyotypes and variant chromosomes thereof

The blue signal is DAPI staining; the green signal is FAM modified oligonucleotide probes pSc119.2-1 and (GAA)₁₀(ii) a The red signal is TAMRA modified oligonucleotide probes pAs1-1, pAs1-3, pAs1-4, pAs1-6, AFA-3 and AFA-4. a: y158; b: FSXM; c: comparison of Y158 with FSXM karyotype; the red boxes represent polymorphic chromosomes present between parents; arrowFirst shown polymorphic chromosome segment

FIG. 2: Y158/FSXM F₂Individual plant FISH with the same color as figure 1

FIG. 3: genetic differentiation index analysis based on wheat 15K chip chromosome polymorphism types PerInv6B and 6B-1

The position of the marker on the chromosome is marked by the horizontal coordinate, the unit is bp, and the Fst value is marked by the vertical coordinate

FIG. 4: genetic differentiation index analysis based on wheat 15K chip chromosome polymorphism types 5A-1 and 5A-4

The position of the marker on the chromosome is marked by the horizontal coordinate, the unit is bp, and the Fst value is marked by the vertical coordinate

FIG. 5: GO annotation of genes within 6B difference segments

FIG. 6: GO annotation of genes within 5A difference segments

FIG. 7: t-test analysis of trait between PerInv6B and class 6B clusters

Indicates significant difference, indicates very significant difference, and NS indicates insignificant difference; a: nanjing; b: linfen

FIG. 8: t-test analysis of traits between 4A-1 and 4A-3 clusters

Indicates significant difference, indicates very significant difference, and NS indicates insignificant difference; a: nanjing; b: linfen

FIG. 9: trait T-test analysis between 5A-1 and 5A-4 clusters

Indicates significant difference, indicates very significant difference, and NS indicates insignificant difference; a: nanjing; b: linfen

FIG. 10: t-test analysis of traits between 4B-1 and 4B-2 groups

Indicates significant difference, indicates very significant difference, and NS indicates insignificant difference; a: nanjing; b: linfen

Fifth, detailed description of the invention

1. Identification of variant chromosomes in Yangmai 158 and Fangshan wheat

The oligonucleotide probe set developed by Du et al.2017 (see references Du P, Zhuang L, Wang Y, Yuan L, Wang Q, Wang D, Dawadondup, Tan L, Shen J, Xu H, Zhuao H, Chu C, Qi Z.development of oligonucleotides and multiplex probes for purposes and acquisition identification of wheat and thin beer Bessarabics chromosomes [ J ]. Genome, 2017, 60: 93-103) was used for the analysis of wheat varieties modified by Huang J, Cheng X, Wang D, Chen J, Bao Y, Guo J, Huang Y, Wang J, Huang H, Zhu H, Chu J # 1, Huang J, Huang D, Cheng J, wheat varieties of wheat, wheat varieties # modified by Huang H, Shu J, Huang J, Qian H, Huang H, and Huang H # 2, and chromosome # 2, wheat varieties, wheat. According to the nomenclature of polymorphic chromosomes in Huang et al (2018), Y158-4A, Y158-5A, Y158-4B, Y158-6B, Y158-7B corresponds to 4A-3, 5A-4, 4B-2, perInv6B and 7B-1 respectively; FSXM-4A, FSXM-5A, FSXM-4B, FSXM-6B, FSXM-7B corresponds to 4A-1, 5A-1, 4B-1, 6B-1 and 7B-7 respectively. Five pairs of chromosomes are found to have difference between two parents, and 1 chromosome structure variation and 5 pairs of polymorphic chromosomes are involved, and are respectively as follows: 4A-3 vs 4A-1, 5A-4 vs 5A-1, 4B-2 vs 4B-1, PerInv6B vs 6B-1, 7B-1 vs 7B-7 (FIG. 1). 4A-1, the end of the long arm has more green signals than 4A-3; 5A-4 has more than 5A-1 long arm middle part a pair of weak green signals, 5A-1 red signal is obviously stronger than 5A-4; 4B-1 and 4B-2 differ in the presence or absence of a green spot at the proximal end of the short arm, 4B-1 is absent, and 4B-2 is present; 6B differ by the presence or absence of PerInv 6B; the end of the long arm of 7B-1 has a significantly stronger green signal than that of 7B-7.

2. Genetic transmission of variant chromosomes

To understand the polymorphic chromosomes 4A, 5A, 4B, 6B, 7B at Y158/FSXM hybrid F₂Distribution in (1) for 288F strains₂Identification was performed (FIG. 2) and a total of 145 karyotypes were found, wherein the number of Y158 type homozygoses, the number of heterozygous type homozygotes, and the number of FSXM type homozygotes are shown in Table 1. Proved by chi-square test, the ratio of the Y158-4A homozygous type, the Y heterozygous type and the FSXM-4A homozygous type is obviously different from the theoretical separation ratio of 1: 2: 1 (df is 2, chi)²More than 5.99, P is less than 0.05); the Y158-perInv6B homozygous type, parental mixed type, and FSXM-6B homozygous type ratios were significantly different from the theoretical separation ratio of 1: 2: 1 (df 2, χ)²More than 9.21, P is less than 0.01); the remaining polymorphic chromosomes and theoryNo significant difference in separation ratios (df 2, χ)²< 5.99, P > 0.05), indicating normal delivery.

For Y158/FSXM RIL-F₈The identification of 62 families showed that: only the distribution of chromosomes perInv6B and 6B-1 is biased apart (df is 1, χ)²> 3.84, P < 0.05) (Table 1), other chromosomes were isolated normally.

For RIL F₈124 seeds of the self-bred generation of the hybrid PerInv6B individual plant are identified, and the homozygous type, heterozygous type and 6B-1 homozygous type of the PerInv6B are 31: 67: 26, which are consistent with 1: 2: 1.

TABLE 1 polymorphic chromosomes in Yang158/Fangshan wheat F₂And F₈Distribution in

3. Effect of variant chromosomes on population differentiation

For the above identified Y158/FSXM RIL-F₈Genotype analysis of wheat 15K chip is carried out, and genetic differentiation index analysis (Fst) is carried out on five pairs of clusters divided according to wheat variant chromosome types, so that difference SNP intervals exist near corresponding inversion or polymorphism segments between two pairs of clusters of PerInv6B vs 6B-1 (figure 3) and 5A-4 vs 5A-1 (figure 4), and the difference segments respectively

TABLE 2 polymorphic chromosomal Difference segment information

367.05Mb, 51.97Mb (Table 2), the sites of variation among other variant chromosome clusters are less or need to be confirmed further with a higher density of markers. The gene annotation shows that the genes in the difference interval of PerInv6B vs 6B-1 are significantly enriched in 10 functions, mainly related to cell development, cell surface receptor signal transduction pathways and the like (figure 5); the genes in the 5A-4 vs 5A-1 difference interval are significantly enriched in molecular functions such as beta-glucosidase activity, acylglycerol O-acyltransferase activity, etc. (FIG. 6).

4. Effect of variant chromosomes on Gene expression

For Y158/FSXM RIL-F₁₀BSR-seq analysis based on chromosome variation types revealed that SNP differences exist in five regions of the 6B chromosome between PerInv6B and 6B-1 pool, and the SNP differences comprise 801 genes in total, wherein 21 genes are significantly differentially expressed between parents and pool at the same time, and alpha-amylase, serine, threonine protein kinase, peroxidase and the like are involved (Table 3).

TABLE 3 Gene annotation of differentially genes in regions of significant 6B differences between pools

5. Influence of variant chromosomes on phenotype

Using Y158/FSXM RIL-F₁₀Two-point three-repeat data analysis of differences between five pairs of variant groups shows that the effects of different variant chromosomes exist in different environments, wherein perInv6B has the effects of reducing plant height in Nanjing, reducing effective spikelet number and panicle number in two places in Nanjing Zheng (FIG. 7), 4A-3 has the effects of reducing plant height, reducing tillering, reducing panicle length and reducing panicle number in Zheng (FIG. 8), 5A-4 has the effects of reducing plant height and shortening panicle length in Zheng (FIG. 9), 4B-2 has the effects of reducing plant height, increasing panicle number and reducing panicle length in Nanjing (FIG. 10), and the obvious phenotypic effect of the variant chromosomes is confirmed, but is influenced by various factors, is not completely consistent in two places, and further repetition is needed.

The above embodiments are merely exemplary embodiments adopted to illustrate the system disclosed in the patent, however, the present invention is not limited thereto, and those skilled in the art can make various modifications and changes without departing from the spirit of the present invention, and these modifications and changes also fall into the scope of the present invention.