基于简化基因组测序筛选白斑狗鱼耐热性状关联的InDel标记

Screening of InDel markers associated with heat tolerance traits in Esox lucius based on simplified genome sequencing

为了能给白斑狗鱼耐高温性状的改良提供有效的分子标记,本研究基于白斑狗鱼热敏感组(109尾)和耐高温组(103尾)进行简化基因组测序,对25个染色体中的InDel标记,以前两个PCA为协变量,利用MLM模型(Masked Language Mode)与白斑狗鱼的耐热性状进行了关联分析。结果显示,大部分InDel分布在内含子(63.69%),外显子分布的InDel位点较少(1.30%)。通过GWAS分析,发现5个位点与白斑狗鱼耐热性状显著关联,分别位于safb基因、未知基因LOC117593903和CLSTN2基因内含子中。其中9N del、4N del-1和4N del-2三个InDel位点均位于CLSTN2基因第3内含子,这3个位点在212尾个体中基因型分布高度连锁。本研究中发现的5个InDel突变可能会对白斑狗鱼的耐热性状产生显著的影响,可作为白斑狗鱼耐热性状改良的候选分子标记。进一步在验证群体中利用KASP技术对部分位点进行了验证。发现9N del位点DD基因型个体在热敏感组中占优势,DI基因型个体在耐高温组中占优势,与简化基因组测序结果基本一致。位于CLSTN2基因第2内含子的3个InDel位点可能影响CLSTN2基因的转录,该基因可能是白斑狗鱼耐高温性状相关的重要候选基因。研究结果为白斑狗鱼分子标记辅助育种提供了理论依据,为白斑狗鱼耐热性状的改良提供了候选分子标记。

To provide effective molecular markers for the improvement of high temperature tolerance of Esox lucius,simplified genome sequencing was conducted based on the heat sensitive group(109)and high temperature tolerant group(103)of E.lucius.For InDel markers in 25 chromosomes,the first two PCA were covariables.The Masked Language Mode(MLM)was used to analyze the correlation between the heat tolerance characteristics of E.Lucius.The results showed that most InDel was distributed in introns(63.69%),while exons had few InDel sites(1.30%).Five loci were found to be significantly associated with heat tolerance traits(FDR P-value<0.05),which were located in the intron of safb gene,unknown gene LOC117593903 and CLSTN2 gene,respectively.The three InDel sites of 9N del,4N del-1 and 4N del-2 were all located in the 3rd intron of Clstn2 gene,and the genotypes of the three sites were highly linked in 212 individuals.The five InDel mutations found in this study may have a sig-nificant effect on the heat tolerance traits of E.lucius,and can be used as candidate molecular markers for the improvement of heat tolerance traits of E.lucius.Further,some loci were verified by KASP technique in valida-tion population.It was found that the 9N del site DD genotype was dominant in the heat sensitive group,and the DI genotype was dominant in the high temperature tolerance group,which was basically consistent with the sim-plified genome sequencing results.Three InDel sites located in the second intron of CLSTN2 gene may affect the transcription of CLSTN2 gene.This gene may be an important candidate gene related to high temperature toler-ance in E.lucius.The results provide theoretical basis for marker assisted breeding and candidate molecular mark-ers for the improvement of E.lucius heat tolerance traits.