利用BSA法定位大豆全基因组株高QTL及候选基因分析

Genome-wide Mapping of QTLs and Candidate Genes Underlying Plant Height in Soybean Using BSA Method

株高是影响大豆单株产量的重要性状,明确调控大豆株高的主效基因对利用分子辅助育种手段提高大豆产量具有重要意义。选用株高差异显著的东农594(高秆)为父本,Charleston(矮秆)为母本,杂交构建F2群体和RIL群体,然后选取F2群体的1500个单株中极高和极矮单株各40株和RIL群体的高矮秆单株各20株,分别建立2套高矮秆池;进而利用二代DNA测序技术对亲本进行50×测序,高矮秆池进行20×测序并筛选read读数大于4的高质量可信SNP位点,在F2群体筛选到20187个符合要求的SNP位点、RIL群体筛选到20285个符合要求的SNP位点;最后采用混合群体分离分析(Bulked segregant analysis,BSA)的SNP-index关联分析法和Euclidean distance(ED)关联分析法计算2个群体和株高关联的染色体区间,进而重新计算株高关联染色体的阈值,并筛选2个群体2种方法共同定位到的10个染色体区间作为候选区间,此结果显著降低了全基因组单一阈值鉴定到的染色体区间,提高了定位结果的准确性。最后利用拟南芥和大豆基因组注释信息对株高关联候选区间的SNP位点所对应的基因功能进行分析,共筛选到18个控制株高的候选基因(Glyma.01G214300、Glyma.01G220700、Glyma.02G095600、Glyma.03G158100、Glyma.03g230000、Glyma.04G033800、Glyma.04G063800、Glyma.04g065600、Glyma.04G190800、Glyma.04G210000、Glyma.04G210700、Glyma.04G212900、Glyma.04g227700、Glyma.05G148700、Glyma.09G158400、Glyma.11G100400、Glyma.12G012400、Glyma.15g052400)。结合大豆旺盛生长期(R1期)顶芽的转录组数据筛选控制大豆株高的重要候选基因,Glyma.03g230000、Glyma.04g065600、Glyma.04g227700在大豆株高形成的关键时期R1期高表达,说明这些候选基因和株高发育密切相关,但是还需要遗传转化验证功能。

Plant height is the key factor for high yield in soybean and the elucidation of the genes underlying plant height can facilitate the breeding of new soybean varieties using molecular assisted selection.In current study,two populations,F2 and recombinant inbred lines(RILs)were constructed by crossing Dongnong 594 and Charleston.40 individuals with extremely high plant heights and another 40 individuals with extremely short plant heights from the 1500 F2∶3 individuals were selected to establish tall and low pools for DNA sequencing.Meantime,20 extremely high individuals and another 20 short individuals from the RIL population were also selected to construct another tall and low pools for DNA sequencing.The parents were sequenced by 50×depth and high and low pools were sequenced by 20×depth,respectively.A total of 20187 reliable SNPs in F2 population and 20285 reliable SNPs in RIL population(reads more than 4)were identified,respectively.The high-quality and reliable SNPs were chosen to identify the candidate genes underlying plant height in soybean using SNP-index association analysis method and Euclidean distance(ED)association analysis method.Ten chromosome segments were identified using F2 population and RIL population with SNP-index and ED method,simultaneously,which improved the mining results from only one population with one calculation method.Further,the annotation information of Arabidopsis thaliana and soybean were used to screen the candidate genes underling the height of soybean.Meantime,we compared the candidate genes with the transcriptome data(from R1 period of tall and low lines of RILs)to further understand the expression way.Finally,18 genes controlling plant height were located on ten different chromosomes of soybean(Glyma.01G214300,Glyma.01G220700,Glyma.02G095600,Glyma.03G158100,Glyma.03g230000,Glyma.04G033800,Glyma.04G063800,Glyma.04g065600,Glyma.04G190800,Glyma.04G210000,Glyma.04G210700,Glyma.04G212900,Glyma.04g227700,Glyma.05G148700,Glyma.09G158400,Glyma.11G100400,Glyma.12G012400 and Glyma.15g052400).Among them,Glyma.03g230000,Glyma.04g065600,and Glyma.04g227700 were detected in the transcriptome data,indicating that these candidate genes are closely related to plant height development,and the function of these genes are studied.Furthermore,the function verification of transgene soybean is needed.It is of great significance to soybean plant assisted breeding.