四倍体马铃薯抗卷叶病毒的分子标记开发及性状关联分析  

Molecular Marker Development and Trait Association Analysis of Resistant to PLRV in Tetraploid Potato

作　　者：杨祝强 梁静思 段晓艳[1,2] 李灿辉[1,2] 齐凤娜 谷爱新 杨洋 唐唯[1,2] Yang Zhuqiang;Liang Jingsi;Duan Xiaoyan;Li Canhui;Qi Fengna;Gu Aixin;Yang Yang;Tang Wei(School of Life Science,Yunnan Normal University,Kunming,650500;Key Laboratory for Potato Biology of Yunnan Province,Joint Academy of Potato Science,Yunnan Normal University,Kunming,650500)

机构地区：[1]云南师范大学生命科学学院,昆明650500 [2]云南师范大学,马铃薯科学研究院,云南省马铃薯生物学重点实验室,昆明650500

出　　处：《分子植物育种》2020年第9期2948-2956,共9页Molecular Plant Breeding

基　　金：国家自然科学基金项目(31660503)资助。

摘　　要：病毒病是马铃薯生产中的主要病害之一,侵染马铃薯的病毒中,影响较为严重和发生较为普遍是卷叶病毒(potato leaf roll virus,PLRV)引起的卷叶病。本研究通过田间性状调查得到50株马铃薯‘合作88’自交F2代抗PLRV性状分离群体,首先利用DAS-ELISA明确侵染病毒的类型,随机选取5株抗病群体和5株感病群体,通过Illumina 2X覆盖度重测序,随后以马铃薯双单倍体DM为参考基因组进行单倍体组装和注释。通过基于Perl语言下的MISA程序进行全基因组SSR挖掘和比对,共筛选出277个与抗卷叶病相关的特异性SSR标记;接下来随机选取180个SSR标记并设计引物,PCR扩增后通过LabChip GX Touch检测得到232个等位位点,其中多态性位点152个,基于Minimum-Evolution的聚类分析发现,在遗传相似性系数为0.80时,能把抗PLRV两个性状分离群体区分。利用JoinMap 4.0构建了‘合作88’抗PLRV连锁的遗传图谱;遗传图谱共包含8个连锁群,总长度为2684.3 cM,标记间平均距离11.57 cM。最后用TetraploidMap检测到5个与抗卷叶病相关的QTL,其遗传贡献率分别为4.31%、8.70%、10.89%、13.52%、7.82%。基于mapping的单倍体测序数据,5个QTL分别被定位于第Ⅹ号、Ⅲ号、Ⅰ号、Ⅷ号、Ⅸ号染色体。本研究可为高通量开发马铃薯抗卷叶病分子标记提供技术参考,也可为精细定位马铃薯优良性状相关基因提供理论依据。Virus disease is one of the most serious diseases in potato industry,especially Leaf roll caused by potato leaf roll virus(PLRV),is more common took place in potato planting areas.In this research,we selected 50 single plants linked to PLRV resistant trait in tetraploid potato’cooperation 88’self-cross F2 generation by a field survey.First using DAS-ELISA to identify the type of infested virus,5 resistant populations and 5 sensitive populations were selected randomly,and 2X deep re-sequencing by Illumina,then haploid assembly and annotation by potato reference genome doubled-monoploid(DM).Besides,the 277 specific SSR locus were identified using MISA program based on Perl environment based on genomic scanning filtering and aligning.Using designed 180 SSR primers,232 alleles were amplified by LabChip GX Touch analyzer among populations,which 152 were polymorphic sites.The clustering analysis based on Minimum-Evolution method,found that when the genetic similarity coefficient was 0.80,the two resistant PLRV traits can be clear classified.The genetic map was constructed using JoinMap 4.0 contained 8 linkage groups with a total length of 2684.3 cM and an average distance between markers was 11.57 cM.Finally,five QTL were detected by TetraploidMap,the genetic contribution rates were 4.31%,8.70%,10.89%,13.52%and 7.82%,respectively.Based on the mapped haploid sequencing data,the five QTL were mapped to chromosomesⅩ,Ⅲ,Ⅰ,Ⅷ,andⅨ.This study can provide a technical reference to the high-throughput development of molecular markers for potato resistant to leaf rolling virus,and also can provide a theoretical basis for gene fine mapping related to potato superior traits.

关 键 词：马铃薯卷叶病毒 SSR 单倍体组装 遗传图谱 QTL 

分 类 号：S435.32[农业科学—农业昆虫与害虫防治]