基于BSA-Seq的家蚕NC99R抗BmNPV分子标记鉴定  

Identification of Anti-BmNPV molecular marker of silkworm NC99R based on BSA-Seq

作　　者：唐亮 蒋满贵 唐名艳 黄深惠 陈小青 董桂清 王平阳 潘志新 TANG Liang;JIANG Man-gui;TANG Ming-yan;HUANG Shen-hui;CHEN Xiao-qing;DONG Gui-qing;WANG Ping-yang;PAN Zhi-xin(Sericulture Technology Promotion Station of Guangxi/Guangxi Academy of Sericulture Sciences,Nanning,Guangxi 530007,China)

机构地区：[1]广西蚕业技术推广站/广西蚕业科学研究院,广西南宁530007

出　　处：《南方农业学报》2023年第8期2406-2414,共9页Journal of Southern Agriculture

基　　金：广西科技重大专项(桂科AA19182012-4);广西重点研发计划项目(桂科AB20297044);国家蚕桑产业技术体系建设专项(CARS-18)。

摘　　要：【目的】以BSA-Seq测序分析确定抗家蚕血液型脓病品种桂蚕N2抗性亲本NC99R的抗性SNP标记及抗性基因,为NC99R抗性基因鉴定打下基础,也为家蚕抗血液型脓病分子标记辅助育种提供标记资源。【方法】以抗病品种NC99R、感病品种芙蓉为亲本制备杂交F1、F2代及BC1遗传群体,通过添食核型多角体病毒(BmNPV)分析其遗传规律;在NC99R近等位基因系NC99R-NIL和芙蓉制备杂交的F2代群体中,设病毒浓度差异达4000倍的高浓度和低浓度添食组,在高浓度添食组挑选未发病个体用于构建极端抗性素材DNA混合池,在低浓度添食组挑选具有典型发病症状个体构建极端易感素材DNA混合池,利用二代测序技术完成抗性DNA混合池、易感DNA混合池及2个亲本基因组测序,并通过群体分离分析法(BSA)定位抗性连锁关联区域。【结果】NC99R对BmNPV表现出稳定的高抗性,基因组中携带有抗BmNPV基因,且由显著的抗性主效基因或紧密连锁基因共同调控。对极端抗性DNA混合池、极端易感DNA混合池及其亲本DNA进行全基因组测序分析,共获得19276670个SNP位点和4789615个InDel位点;基于ΔSNPindex的LOESS回归拟合分析,定位到2个与血液型脓病抗性关联的区域,分别为Chr.25:150-11069kb和Chr.27:8700-11009kb。同时将ΔSNP-index设定为≥0.8时,鉴定到3865个与抗性显著相关的SNP标记,其中,在25号染色体关联区域筛选出78个SNP标记,在27号染色体关联区域筛选出2694个SNP标记;再经氨基酸水平非同义突变筛查,定位到25号染色体Zinc carboxypeptidase基因和27号染色体Facilitated trehalose transporter Tret1基因。【结论】利用BSA-Seq在抗家蚕血液型脓病品种NC99R中发现与抗性相关的SNP标记资源有3865个,且初步鉴定到2个与抗BmNPV的相关基因(25号染色体Zinc carboxypeptidase基因和27号染色体Facilitated trehalose transporter Tret1基因),为家蚕抗血液型脓病分子标记辅助育种提供了标�【Objective】By using BSA-Seq sequencing analysis to determine the resistance SNP markers and resistance genes of the resistant parent NC99R of the nuclear polyhedrosis resistant variety Guican N2,lay a foundation for the identification of NC99R resistance genes and provide marker resources for silkworm nuclear polyhedrosis resistant molecular marker assisted breeding.【Method】A hybrid F1,F2,and BC1 genetic population were prepared using the disease resistant variety NC99R and the susceptible variety Furong as parents,and their genetic patterns were analyzed by feeding nuclear polyhedrosis virus(BmNPV).In the F2 population of NC99R near allelic gene line NC99R-NIL hybridized with Furong,high and low concentration feeding groups with virus concentration differences of 4000 times were set up.Non-diseased individuals were selected from the high concentration feeding group to construct an extreme resistance material DNA mixing pool.Individuals with typical disease symptoms were selected from the low concentration feeding group to construct an extreme susceptibility material DNA mixing pool.Then the resistance DNA mixing pool,susceptible DNA mixed pool and two parental genomes were completed sequencing using second-generation sequencing,and locating resistance linked regions with population segregation analysis(BSA).【Result】The results showed that NC99R exhibited stable high resistance to BmNPV,and the genome carried anti BmNPV genes,which were jointly regulated by significant resistance major genes or closely linked genes.A total of 19276670 SNP loci and 4789615 InDel loci were obtained through whole genome sequencing analysis of extreme resistance DNA mixing pool,extreme susceptibility DNA mixing pool and parental DNA.Based on the LOESS regression fitting analysis ofΔSNP-index,two regions associated with resistance to nuclear polyhedrosis were identified,Chr.25:150-11069kb and Chr.27:8700-11009kb respectively.When theΔSNP-index was set to≥0.8,a total of 3865 SNP markers significantly related to resistan

关 键 词：家蚕 NC99R 核型多角体病毒(BmNPV) 抗性分子标记 BSA-Seq 

分 类 号：S884.51[农业科学—特种经济动物饲养]