中国农业科学

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最新录用:马铃薯品种(系)田间晚疫病抗性评价和全基因组遗传多样性分析

李晓川,王朝海,周平,马维,吴瑞,宋治豪,梅艳   

  1. 毕节市农业科学研究所,贵州毕节 551700
  • 发布日期:2022-08-19

Deciphering of the Genetic Diversity After Field Late Blight Resistance Evaluation of Potato Breeds

LI XiaoChuan, WANG ChaoHai, ZHOU Ping, MA Wei, WU Rui, SONG ZhiHao, MEI Yan   

  1. Bijie institute of agriculture science, Bijie 551700, Guizhou
  • Online:2022-08-19

摘要: 【目的】进行田间晚疫病抗性评价,利用SNP分子标记分析马铃薯抗晚疫病种质的遗传多样性及分离基因组内可能影响马铃薯晚疫病抗病性状的遗传区段,为马铃薯晚疫病抗性种质创新与利用提供理论依据。【方法】通过多年多点田间鉴定对马铃薯种质资源进行晚疫病抗性评价。利用dd-RAD技术对马铃薯种质资源进行简化基因组测序并分型SNP标记。利用Admixture软件分析群体的遗传结构,GCTA软件进行主成分分析,fastTree软件构建系统发育树,stacks程序包中的populations命令计算群体遗传多样性参数的,vcftools程序计算选择性消除参数Clustal Omega程序比对氨基酸序列MEGA6绘制氨基酸序列进化树,GEMMA 0.98.1软件进行全基因组关联分析,CMplot程序绘制QQ图和曼哈顿图。【结果】通过对马铃薯种质资源多年多点田间晚疫病抗性鉴定,得到101个抗晚疫病的品种(系)21个感病品种并对它们进行dd-RAD简化基因组测序,通过对比参考基因组,共检测到分布相对均匀的8 697 602SNP。通过种群结构分析、主成分分析和系统进化分析可以将这些种质资源进一步划分为6个群体。6个群体内的平均核苷酸多样性值π为0.2055—0.2572间的群体分化指数Fst为0.156909—0.187336,说明这些种质资源存在较丰富的单核苷酸多态性。同时6个群体内期望杂合度He为0.187—0.2297观测杂合度(Ho为0.0829—0.11866个群体内的观测杂合度均小于期望杂合度,并且6个群体内的近交系数(Fis范围为0.2412—0.3554,说明在选育这些种质的过程中存在近交现象。分析可能影响晚疫病抗性的马铃薯基因组遗传区段,20 kb为窗口5 kb为步长,在基因组相同位置,分别计算不同抗性种质间π值比值和Fst值,进行选择性消除分析,选择π值比值最小的5%Fst值最大的10%745个遗传区段进行分析,遗传区段中共包含507个基因,其中4NBS-LRR类基因。利用群体SNP和马铃薯种质的不同晚疫病抗性表型,进行全基因组关联分析,显示有9SNP与抗病性状高度相关,周围50 kb的基因组范围内,有69个基因,其中15个基因预测参与到应激反应,12个基因预测参与清除过氧化物自由基过程。【结论】dd-RAD简化基因组测序可以在马铃薯基因组中分型到数量较多分布相对均匀的SNP标记。马铃薯田间抗晚疫病种质资源拥有较丰富的单核苷酸多态性但在其选育过程中存在近交现象。选择性清除和关联分析有助于分离影响晚疫病抗病性状的遗传区段。

关键词: 马铃薯, 晚疫病, 单核苷酸多态性标记, 遗传多样性, 选择性消除, 关联分析

Abstract: 【ObjectiveTo evaluate the late blight resistance in field. To use SNP markers to analyze the genetic diversity of late blight resistance of potato germplasm and discern the genetic segments that may affect the phenotype of potato late blight resistance, and to provide a theoretical basis for the innovation and utilization of potato late blight resistant germplasm.MethodField resistance to late blight was evaluated in potato germplasm at multiple locations and over the course of several years. SNP markers were detected using a dd-RAD simplified genome sequencing strategy. The population genetic structure was analyzed using Admixture, principal component analysis was conducted using GCTA, a phylogenetic tree was constructed using fastTree, population genetic diversity parameters were calculated using the populations command in the Stacks package, selective sweep parameters were calculated using vcftools, protein sequences were aligned using Clustal Omega, and a proteins phylogenetic tree was drawn using MEGA6. Genome-wide association analysis was constructed using GEMMA 0.98.1 and QQ and manhattan plots were drawn using CMplot.ResultThrough years of evaluation of late blight field resistance in multiple locations, potato germplasm of 101 late blight resistant varieties (lines) and 21 susceptible varieties were obtained. A total of 8 697 602 relatively evenly distributed SNPs were obtained using dd-RAD simplified genome sequencing on these germplasms. This germplasm can be further divided into 6 populations through structural analysis, principal component analysis and phylogenetic analysis. The average nucleotide diversity (π) within the 6 populations ranged from 0.2055 to 0.2572 and the fixation index (Fst) among the six populations ranged from 0.156909 to 0.187336, revealing a relatively large genetic diversity for these germplasms. The expected heterozygosity (He) within the 6 populations ranged from 0.187 to 0.2297 and the observed heterozygosity (Ho) ranged from 0.0829 to 0.1186. The values of Ho were less than those of He in all six populations. Meanwhile, the inbreeding coefficient (Fis) for the six populations ranged from 0.2412 to 0.3554, indicating inbreeding events during the breeding process. To identify the genetic segments that may affect the phenotype of potato late blight resistance, π ratios and Fst among different late blight resistance germplasms in the whole potato genome were calculated using 20 kb as the window length and 5 kb as the step length. The 745 genetic segments which had a π ratio value in the lowest 5% and a Fst value in highest 10% were further analyzed by performing a selective sweep analysis. These selected segments contain a total of 507 genes, including 4 NBS-LRR genes. A genome-wide association analysis was also conducted, yielding 9 SNP highly associated with late blight resistance. Of the 69 genes located in the genome within 50 kb around the 9 SNPs, 15 genes were predicted to be involved in stress response, and 12 genes were predicted to be involved in removing peroxide radicals.ConclusionLarge amount of SNPs, which are relatively evenly distributed in the potato genome, can be genotyped by dd-RAD simplified genome sequencing. Potato late blight field resistance germplasm has a large genetic diversity, but had inbreeding events in the process of breeding. Population structure analysis can reveal the genetic relationship between potato germplasms, which can further provide a theoretical basis for parental selection in breeding. Selective sweep and genome-wide association analyses help to isolate genetic segments that may affect late blight resistance traits.

Key words: potato (Solanum tuberosum L.), late blight (Phytophthora infestans), single nucleotide polymorphism (SNP), genetic diversity, selective sweep, association mapping