80份绿豆象抗感蚕豆种质资源SSR遗传多样性分析

Genetic diversity of SSR markers in 80 faba bean germplasm resources susceptible to mung bean weevil

【目的】通过转录组测序开发的SSR分子标记分析蚕豆种质资源遗传多样性,挖掘与蚕豆抗豆象性状相关的分子标记,为蚕豆抗豆象种质资源的保护利用及新品种选育提供参考依据。【方法】采用PacBio三代全长测序技术结合RNA-seq方法,对2个蚕豆品种Y134和Y078进行转录组测序,在此基础上,使用软件MISA对转录组所有Unigene序列进行搜索,寻找包含SSR核心基序的Unigene序列,并利用Primer 3.0批量设计引物。随机挑选100对SSR引物对80份蚕豆种质资源进行遗传多样性分析。【结果】100对SSR引物中有50对扩增出产物,有效扩增率为50%;其中32对引物表现出多态性。占有效产物的64%,占总引物的32%。在32对多态性引物中,每对引物扩增等位基因2~21个,平均值为6.875个;多态信息量(PIC)值为0.25~0.82,平均值为0.58;Shannon信息指数变化范围为0.34~2.31,平均值为1.21。基于UPGMA聚类分析将80份蚕豆材料在相似系数0.70处分为两大类:第1类包含65份种质,其中高抗(HR)品种62份,高感(HS)品种2份,感(S)品种1份;第2类包含15份种质,其中高感(HS)品种7份,感(S)2份,高抗(HR)品种5份,中抗(MR)1份。【结论】利用开发的SSR标记分析了80份蚕豆种质资源的遗传多样性,为蚕豆抗豆象种质资源鉴定与分子标记辅助选择育种等方面的研究奠定了理论基础。

[Objective]The aim of the paper is to analyze the genetic diversity of faba bean germplasm resources by transcriptome sequencing and SSR molecular marker technology,and to explore the molecular markers associated with bruchid resistance traits in faba bean.This can provide a reference basis for the conservation and utilization of bruchid resistance germplasm resources and the selection and breeding of new varieties.[Method]Transcriptome sequencing of two faba bean varieties Y134 and Y078 was performed by PacBio triple full-length sequencing technology combined with RNA-seq.Based on this,all Unigenes of the transcriptome were searched for Unigene containing SSR core motifs using the software MISA,and primers were designed in batch using Primer 3.0.One hundred pairs of SSR primers were randomly selected for genetic diversity analysis of 80 faba bean germplasm resources.[Result]Fifty of the 100 SSR primer pairs amplified products,with an effective amplification rate of 50%;32 of the primer pairs showed polymorphism.They accounted for 64%of the effective products and 32%of the total primers.Among the 32 polymorphic primer pairs,each pair amplified 2 to 21 alleles with an average of 6.875;The polymorphic information content(PIC)values ranged from 0.25 to 0.82 with an average of 0.58;The Shannon's information index varied from 0.34 to 2.31 with an average of 1.21.Based on UPGMA cluster analysis,the 80 faba bean materials were classified into two major groups at a similarity coefficient of 0.70.The first category contained 65 germplasm,including 62 highly resistant(HR)varieties,2 highly sensitive(HS)varieties and 1 sensitive(S)variety;The second category contained 15 germplasm,7 highly susceptible(HS)varieties,2 sensitive(S)varieties,5 highly resistant(HR)varieties and 1 moderately resistant(MR)variety.[Conclusion]The SSR markers developed were used to analyze the genetic diversity of 80 faba bean germplasm resources and to lay the theoretical foundation for research on the identification of faba bean germplasm resources resistant to bean weevil and molecular marker-assisted selection for breeding.