低温胁迫下超甜玉米种子发芽率全基因组关联分析及关键基因挖掘

Genome-wide association study on super sweet maize seed germination rate and key gene identification under low temperature stress

  • 摘要: 【目的】揭示低温胁迫下超甜玉米种子发芽性状的遗传特性,挖掘与其显著关联的单核苷酸多态性位点(SNP)并筛选关键基因,为探究超甜玉米种子萌发遗传机制及选育高萌发率的超甜玉米品种提供参考依据。【方法】以100个超甜玉米自交系为研究对象,分析低温胁迫下种子发芽率表型性状。利用玉米56K基因芯片对超甜玉米种子低温发芽率进行全基因组关联分析(GWAS),定位低温胁迫下调控种子发芽率主要SNP位点,结合MaizeGDB数据库提取上、下游200 kb区间中的候选基因,对候选基因进行GO功能注释及KEGG信号通路富集分析,并通过分析低温胁迫下甜玉米种子组织中候选基因平均表达量筛选出关键基因。【结果】低温胁迫下超甜玉米种子的发芽率为8%~52%,平均发芽率为20%,标准差为0.073,偏度为1.967,峰度为7.032,变异系数为35.7%。低温胁迫下共关联定位到6个与超甜玉米种子低温发芽率显著关联的SNPs位点,贡献率为22.9%~28.0%,其中贡献率最高是Affx-90570229位点。SNP位点中,Affx-90621601和Affx-91200146与相邻位点呈高度连锁状态,且T/T、C/C、A/A这3个优异等位变异均具有提高种子低温发芽率的特性。基于显著SNP位点,并结合MaizeGDB等数据库共得到103个与种子发芽率相关的候选基因,其中已成功注释的候选基因有21个,占比为20.4%,分别富集到24条GO功能条目中。21个候选基因共富集在12条信号通路中。基因表达差异分析筛选出液泡分选受体(GRMZM2G303631)、四三肽重复序列(TPR)样超家族蛋白(GRMZM2G071986)、HNH核酸内切酶结构域的蛋白(GRMZM2G074804)、MicroRNA(GRMZM2G122199)、微管蛋白折叠辅助因子D(GRMZM5G816015)5个调控低温胁迫下种子发芽率的关键基因,其中GRMZM2G303631基因的平均表达量最高,为165.72。【结论】关联定位到6个控制超甜玉米种子低温发芽率的显著SNPs位点,筛选到5个调控超甜玉米种子低温发芽率相关的关键基因,主要参与酶和蛋白质合成及信号传导等过程。

     

    Abstract: 【Objective】To reveal the genetic characteristics of germination traits in super sweet maize seeds under low temperature stress, identify significant single nucleotide polymorphism sites(SNP) associated with these traits and screen key genes, which could provide reference for exploring the genetic mechanisms of seed germination in super sweet maize and breeding high-germination-rate super sweet maize varieties. 【Method】Using 100 samples of super sweet maize inbred lines as the research materials, the phenotypic traits of seed germination rate under low temperature stress were analyzed. A maize 56K SNP chip was employed to conduct a genome-wide association study(GWAS) on the low-temperature germination rate of super sweet maize seeds, identifying major SNP loci associated with germination rate regulation under low temperature stress. Based on the MaizeGDB database, candidate genes within 200 kb upstream and downstream regions were extracted. GO functional annotation and KEGG signaling pathway enrichment analysis were performed on these candidate genes. Additionally, key genes were screened from the candidate genes by analyzing their average expression levels in sweet maize seed tissues under low-temperature stress. 【Result】Under low temperature stress, the germination rate of super sweet maize seeds ranged from 8% to 52%, with an average germination rate of 20%, a standard deviation of 0.073, a skewness of 1.967, a kurtosis of 7.032, and a coefficient of variation of 35.7%. A total of 6 SNPs were significantly associated with the low-temperature germination rate of super sweet maize seeds under low temperature stress, with a contribution rate of 22.9% to 28.0%. Among them, the SNP locus with the highest contribution rate was Affx-90570229. Among the SNP loci, Affx-90621601 and Affx-91200146 were in a highly linked state with adjacent loci, and the 3 superior allelic variations of T/T, C/C and A/A all had the characteristic of improving the low-temperature germination rate of seeds.Based on the significant SNP loci and combined with databases such as MaizeGDB, a total of 103 candidate genes related to seed germination rate were obtained, among which 21 candidate genes were successfully annotated, accounting for 20.4%, and were enriched in 24 GO functional terms. The 21 candidate genes were enriched in 12 signaling pathways. Gene expression difference analysis screened out 5 key genes regulating the germination rate of seeds under low temperature stress: vacuolar sorting receptor(GRMZM2G303631), tetratricopeptide repeat(TPR)-like superfamily protein(GRMZM2G071986),HNH endonuclease domain-containing protein(GRMZM2G074804),microRNA(GRMZM2G 122199) and tubulin folding cofactor D(GRMZM5G816015). Among them, GRMZM2G303631 gene had the highest average expression level(165.72). 【Conclusion】Six significant SNPs loci associated with the lowtemperature germination rate of super sweet maize seeds were identified, and five key genes related to the regulation of the low-temperature germination rate of super sweet maize seeds were screened out, mainly involved in enzyme and protein synthesis and signal transduction processes.

     

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