摘要：

干旱是限制粳稻产量的重要因素之一,探究萌发期粳稻的耐旱性遗传机制,筛选耐旱候选基因,挖掘其优异单倍型,为解析粳稻萌发期耐旱机制及开发相关分子标记提供理论基础。采用西北粳稻核心种质资源139份为材料,15% PEG-6000模拟干旱胁迫露白后的粳稻种子,测定相对发芽率、相对根长等6个耐旱相关指标,并结合255 501个高质量单核苷酸多态性(SNPs),运用一般线性模型(GLM)、混合线性模型(MLM)和固定与随机效应循环概率统一模型(FarmCPU)开展全基因组关联分析(GWAS),并结合耐旱表型数据进行单倍型分析,进而挖掘潜在的候选基因。结果表明,6个耐旱指标均值呈正态分布,且变异系数为3.43%~56.25%,其中以相对根芽干质量变异系数最大,与相对根长、相对芽长、相对根芽鲜质量间均呈显著正相关。结合高密度SNPs,对6个耐旱性状进行全基因组关联分析,共检测到1 164个不同的SNPs,其中379个被视为高可信度SNP位点,6个被视为数量性状核苷酸(QTNs)。整合这些位点的上下游50 kb区间,共筛选出14个显著SNPs位点,对应14个可信的QTL候选区间,且主要与相对根芽干质量显著关联。通过数据库进一步预测分析14个区间内的基因,并以外显子具有错义突变为标准,筛选到了编码地衣酶-2前体的LOC_Os03g56130基因。结合耐旱表型,对该基因进行单倍型分析,发现5个外显子错义突变组成了4种单倍型,其中Hap.2的相对根芽干质量显著高于其他单倍型,为优异单倍型。这些研究表明,LOC_Os03g56130为粳稻萌发期耐旱性相关的潜在候选基因。

关键词: 粳稻, 全基因组关联分析, 耐旱性, 萌发期, 基因挖掘

Abstract:

Drought is one of the key factors limiting the yield of japonica rice.Exploring the genetic mechanism of drought tolerance in japonica rice at the germination stage,screening drought-tolerant candidate genes,and identifying their superior haplotypes provide a theoretical basis for deciphering the drought tolerance mechanism of japonica rice at the germination stage and developing related molecular markers.139 core germplasm resources of japonica rice from Northwest China were used as materials.Japonica rice seeds after radicle emergence were subjected to simulated drought stress using 15% PEG-6000.Six drought tolerance-related traits,including relative germination rate(RGR)and relative root length(RRL),were determined.Combined with 255 501 high-quality single nucleotide polymorphisms(SNPs),genome-wide association studies(GWAS)were conducted using the general linear model(GLM),mixed linear model(MLM),and fixed and random model circulating probability unification(FarmCPU)model.Haplotype analysis was further performed based on drought tolerance phenotypic data to identify potential candidate genes.The results showed that the mean values of the six drought tolerance traits followed a normal distribution,with coefficients of variation(CV)ranging from 3.43% to 56.25%.Among these traits,the relative root and shoot dry weight(RRSDW)exhibited the largest CV and showed a significantly positive correlation with relative root length(RRL),relative shoot length(RSL),and relative root and shoot fresh weight(RRSFW).GWAS for the six drought tolerance traits using high-density SNPs detected a total of 1 164 distinct SNPs,among which 379 were regarded as high-confidence SNPs loci and 6 as quantitative trait nucleotides(QTNs).By integrating the 50 kb upstream and downstream regions of these loci,14 significant SNPs loci were screened out,corresponding to 14 reliable quantitative trait loci(QTL)candidate intervals,which were mainly significantly associated with RRSDW.Genes within these 14 intervals were further predicted and analyzed using databases.With missense mutations in exons as the criterion,the gene LOC_Os03g56130,which encodes lichenase-2 precursor,was identified.Combined with drought tolerance phenotypes,haplotype analysis of this gene revealed that 5 missense mutations in exons constituted 4 haplotypes.Among them,Hap.2 showed a significantly higher RRSDW than the other haplotypes,being identified as the superior haplotype.These findings indicate that LOC_Os03g56130 is a potential candidate gene associated with drought tolerance in japonica rice at the germination stage.

Key words: Japonica rice, Genome-wide association study, Drought tolerance, Germination stage, Gene mining

中图分类号: 

• S511.22

• Q943.2植物基因工程