基于BSA-seq技术对西瓜抗枯萎病生理小种1的基因定位

Mapping of watermelon(Citrullus lanatus)Fusarium wilt-resistance gene Fon-1 based on BSA-Sequencing technology

西瓜枯萎病是由尖孢镰刀菌西瓜专化型(Fusarium oxysporum f.sp.niveum)引起的土传真菌病害,是西瓜生产的限制因子。本研究以西瓜抗病种质“V13-9-3”和感病种质“W16-1”为亲本,构建了“W16-1”דV13-9-3”的F1、F2群体。苗期人工接种结果显示:F1植株表现抗病,F2群体的抗病∶感病分离比为3∶1,表明西瓜枯萎病生理小种1的抗性遗传受显性单基因控制。利用BSA技术,采用SNP指数和InDel指数法,将西瓜抗枯萎病基因Fon-1定位在1号染色体上2.20 Mb和2.32 Mb区域内,将这2种方法的定位结果取交集,最终将目标抗病基因区域缩小在2.20 Mb之内,该区域包含8个非同义突变基因和1个移码突变基因。利用荧光定量PCR检测这9个突变基因在接种枯萎病菌5 d和10 d后的表达量变化,发现抗病亲本“V13-9-3”中有7个基因的表达量显著高于感病亲本“W16-1”。研究通过结合SNP指数和InDel指数分析可以快速进行基因初定位,为西瓜抗枯萎病的分子机理研究提供理论依据。

Watermelon Fusarium wilt,a worldwide soil borne fungal disease,is caused by Fusarium oxysporum f.sp.niveum and can dramatically reduce the watermelon yield.In this study,the resistant watermelon germplasm“V13-9-3”and the susceptible germplasm“W16-1”were used as parents to construct F1 and F2 populations.The F1 population showed resistance to Fusarium wilt,while the F2 population had a 3∶1 segregation ratio of resistance and susceptibility.It demonstrated that the resistance against F.oxysporum f.sp.niveum race 1 was controlled by a single dominant gene.Fon-1 was found to be localized to 2.20 Mb and 2.32 Mb on chromosome 1 by single nucleotide polymorphism(SNP)and insertion deletion(InDel)index methods.After accounting for overlapping regions,the target region narrowed down to 2.20 Mb.Reverse-transcription quantitative polymerase chain reaction analysis further confirmed the relative expression of genes,whereby the expression of 7 genes in“V13-9-3”was significantly higher than that in“W16-1”at 5 dpi or 10 dpi.This study proposed a novel combinatorial strategy by combining the SNP and InDel index analyses.This approach allowed rapid mapping of the genes responsible for specific traits and provides a robust base for cloning the Fon-1 gene.