2个NBS-LRR基因在不同森林草莓生态型中的遗传变异分析  

Genetic variation analysis of two NBS-LRR genes in different woodland strawberry ecotypes

作　　者：仲岩 张甜 陈钊 程宗明[1] ZHONG Yan;ZHANG Tian;CHEN Zhao;CHENG Zongming(College of Horticulture,Nanjing Agricultural University,Nanjing 210095,China)

机构地区：[1]南京农业大学园艺学院,江苏南京210095

出　　处：《南京农业大学学报》2022年第4期666-674,共9页Journal of Nanjing Agricultural University

基　　金：江苏高校优势学科建设工程资助项目。

摘　　要：[目的]本文旨在研究取自欧洲不同生态环境下的森林草莓生态型中的2个NBS-LRR抗病基因,揭示不同生态型中同一基因的遗传变异特征。[方法]选取森林草莓中具有代表性的2个NBS-LRR基因为目的基因,从不同森林草莓生态型中对其进行PCR扩增、测序并得到相应的核苷酸序列。利用ProtParam网站分析其理化性质;利用MEGA v7.0和DNAMAN v7.0对核苷酸序列以及蛋白序列进行多重比对,查找序列突变位点;利用Pfam、SMART、COILS等网站分析基因编码蛋白质的结构域组成;利用MEGA v7.0计算2个基因中不同生态型之间的K_(a)/K_(s)值、Pi值并构建系统进化树,分析其受到的选择压力及遗传进化关系。[结果]以‘Hawaii4’的序列为参照,FvH 4_5g22710和FvH4_1g15230基因在某些森林草莓生态型中发生了点突变和缺失突变。通过结构域分析可知FvH 4_5g22710基因只在Rx_N结构域区段发生了突变,而在NB-ARC结构域和LRR基序中没有发生,FvH 4_1g15230基因在其编码蛋白质的RPW8、NB-ARC结构域和LRR基序区段均发生了突变。进化树的拓扑结构显示,来自在同一国家的生态型,其目的基因更偏向于聚类在同一进化支上;2个基因的平均K_(a)/K_(s)值均小于1,说明它们均受到纯化选择的驱动。[结论]2个NBS-LRR基因在不同的森林草莓生态型中都有一定程度的变异,这提高了抗病基因的遗传变异程度,可能与提高森林草莓抗病性有一定关联。[Objectives]The purpose of this study was to study the variation and evolution of two NBS-LRR genes in different woodland strawberry ecotypes.[Methods]Two representative NBS-LRR resistance genes were selected,and PCR amplification and sequencing were performed to get their nucleotide sequences in different ecotypes.The physicochemical properties were analyzed by using ProtParam website.Multiple alignments for nucleotide sequences and protein sequences were performed using MEGA v7.0 and DNAMAN v7.0 to uncover the mutation sites among the ecotypes.The domains of the proteins were analyzed by using Pfam,SMART,and COILS websites.The K_(a)/K_(s) values,Pi values and the phylogenetic trees of the two genes between different ecotypes were calculated and constructed by MEGA v7.0,and the selection pressure working on them and the evolutionary relationships were analyzed.[Results]It was found that FvH 4_5g22710 and FvH 4_1g15230 gene had point mutations and deletion mutations in some strawberry ecotypes taking the‘Hawaii4’sequences as the references.The results showed that the mutation sites were only found in the Rx_N domain region in the FvH 4_5g22710,but not in NB-ARC domain region and LRR motifs.However,the mutation sites could be detected in RPW8,NB-ARC domain regions and LRR motifs in the FvH 4_1g15230.The evolutionary tree showed that most or all of the strawberry samples from different sites in the same sampling country were distributed on the same branch.The average K_(a)/K_(s) values of the two genes were all less than 1,indicating that they were driven by the purifying selection.[Conclusions]There were mutations in the two NBS-LRR genes between the different ecotypes,which improved the genetic variations of the resistance genes and might be related to the enhancements of strawberry resistance.

关 键 词：森林草莓 生态型 NBS-LRR基因 突变 选择压力 

分 类 号：S668.4[农业科学—果树学]