苦瓜高通量遗传变异检测与枯萎病抗性位点精准鉴定

Fusarium wilt, caused by Fusarium oxyspoorum Schl. f. sp. momodicae Sun & Huang, affects the yield and quality of bitter gourd seriously. Developing resistant bitter gourd cultivars is the most effective and economical method to prevent this disease at present. Crosses were made between bitter gourd line Thai4-6, high resistant to fusarium wilt and CN19-1, high susceptible to fusarium wilt. The inheritance of fusarium wilt resistance will be analyzed with P1, P2, F1, BCP1, BCP2 and F2 populations through employing major gene plus polygene mixed inheritance model..The high throughput SNP and InDel molecular markers of bitter gourd will be developed through GBS (Genotyping by sequencing) technique. Combining the EST-SSR and SSR markers available, the high density genetic linkage map of bitter gourd will be constructed based on the F2 segregating population derived from a cross between high resistant bitter gourd line Thai4-6 and high susceptible line CN19-1. The QTLs of fusarium wilt resistance in bitter gourd will be mapped through integrating the high density genetic linkage map and phenotype data. The molecular markers closely linked with main QTLs loci will be identified. .The study results could provide important information for revealing the genetic mechanism of fusarium wilt resistance and accelerate the breeding process of fusarium wilt resistance.

苦瓜枯萎病是由尖孢镰刀菌苦瓜专化型引起的严重危害苦瓜产量和品质的世界性病害，培育抗病品种是防治苦瓜枯萎病最为经济和有效的方法。本项目利用高抗枯萎病苦瓜种质Thai4-6和高感枯萎病苦瓜种质CN19-1配制杂交组合，采用数量性状主基因+多基因混合遗传模型分析方法对该组合6世代遗传群体（P1、P2、F1、BCP1、BCP2和F2）的枯萎病抗性进行遗传分析，探索抗性主基因与多基因的作用；采用GBS（Genotyping by sequencing）简化基因组测序技术高通量开发SNP、InDel遗传变异，结合已开发的EST-SSR和SSR标记，基于Thai4-6和CN19-1杂交产生的F2分离群体，构建高密度遗传连锁图谱，结合抗病性鉴定数据，准确定位苦瓜抗枯萎病QTLs位点，鉴定与主效QTLs位点紧密连锁的分子标记。研究结果将为解析苦瓜枯萎病抗性遗传机制提供重要信息，加速苦瓜抗枯萎病育种的进程。

以抗枯萎病苦瓜材料Thai4-6和感病材料CN19-1为亲本配制杂交组合，基于该组合6世代遗传群体（P1、P2、F1、F2、BCP1和BCP2），采用主基因+多基因混合遗传模型分析枯萎病抗性遗传特性。数据分析结果表明，该杂交组合的枯萎病抗性呈连续分布，最适模型为2对加性-显性-上位性主基因+加性-显性多基因遗传模型（E-1），2对主基因加性效应值均为-13.85，显性效应值分别是25.58和34.26，主基因遗传率在BCP1、BCP2和F2中分别是86.03%、80.34%和94.25%，表明该组合枯萎病抗性主要受2对主基因控制。基于F2单世代分离群体分析单瓜重和瓜长遗传特性。分析结果表明，该杂交组合的单瓜重和瓜长呈连续分布，最适模型均为1对加性-显性主基因遗传模型（A-1）。.亲本材料CN19-1和Thai4-6基因组重测序分析结果表明，CN19-1共获得高质量reads数为83708504，占99.56%，比对上参考基因组的reads数目为80529117，占96.20%，共鉴定出174819个SNP，40715个InDel。Thai4-6共获得高质量reads数为62288776，占99.62%，比对上参考基因组的reads数目为60318176，占96.84%，共鉴定出592599个SNP，118253个InDel。F2群体GBS测序结果分析表明，高质量碱基所占比例均超过95%，高质量reads所占比例均超过99%，高质量reads与参考基因组的比对率均维持在98%左右，共发现SNP 285258个，InDel 21222个。基于SNP和InDel标记构建的苦瓜高密度遗传图谱共包括11个连锁群，20676个标记位点，全长2727.876 cM，标记间平均距离为0.132 cM。基于区间定位法，苦瓜枯萎病抗性共定位7个QTLs位点，瓜长共定位5个QTLs位点，单瓜重共定位6个QTLs位点。基于复合区间定位法，枯萎病抗性共定位2个QTLs位点，瓜长共定位3个QTLs位点，单瓜重共定位3个QTLs位点。控制枯萎病抗性的QTL位点IM-FW1.3在两种分析方法中均被检测到，对表型的贡献率分别为23.32%和19.25%，表明该位点是稳定的抗枯萎病主效QTL位点，与该位点紧密连锁的标记为cLG02.loc145。