水稻芽期和苗期耐冷性的数量性状位点定位及候选基因表达分析

Quantitative trait locus(QTL) mapping and candidate gene expression analysis of cold tolerance of rice at plumule and seedling stages

摘要:

水稻(Oryza sativa L.)作为一种喜温作物，生长过程中极易受到低温冷害的影响。芽期和苗期低温冷害常影响水稻形态建成并导致减产。耐冷基因的挖掘与利用是解决水稻低温冷害最直接有效的手段之一。为挖掘水稻芽期和苗期耐冷性相关的数量性状位点(quantitative trait locus, QTL)，本研究以籼稻‘华占’(Oryza sativa L. subsp. indica cv. ‘HZ’)、粳稻‘热研2号’(Oryza sativa L. subsp. japonica cv. ‘Nekken2’)及其构建的120个重组自交系(recombinant inbred lines, RILs)为实验材料，测定低温胁迫后水稻芽期成苗率和苗期存活率，基于前期构建的高密度遗传图谱进行水稻芽期和苗期耐冷性相关的QTL定位。结果共检测到4个芽期耐冷性QTLs和9个苗期耐冷性QTLs，其中最大检出限(limit of detection, LOD)值为5.20，并在8号染色体上定位得到1个芽期和苗期遗传重叠的QTL，物理距离为24 432 953–25 295 129 bp。对检测到的QTL区间进行候选基因筛选，并利用实时荧光定量PCR (quantitative real-time polymerase chain reaction, qRT-PCR)进行芽期和苗期基因表达量分析，发现LOC_Os03g06570、LOC_Os03g07100、LOC_Os06g08280、LOC_Os08g38440、LOC_Os08g39100、LOC_Os08g39540在双亲间的表达量差异显著，且在冷胁迫后显著下调或上调表达，推测前3个基因为调控水稻苗期耐冷性的重点候选基因，后3个基因为综合调控水稻芽期与苗期耐冷性的重点候选基因。研究结果为芽期和苗期耐冷基因的精细定位和克隆奠定了基础，对培育耐冷性水稻品种有重要意义。

Abstract:

Rice (Oryza sativa L.), as a thermophilic crop, is highly susceptible to cold stress during its growth process. Chilling injury at the plumule stage and seedling stage often affects the morphological development and leads to yield reduction of rice. The exploration and utilization of cold tolerance genes are among the most direct and effective approaches to address cold stress in rice. To identify quantitative trait loci (QTLs) associated with cold tolerance at plumule and seedling stages, in this study, we measured the seedling rates and survived seedling rates of the indica rice cultivar 'HZ', the japonica cultivar 'Nekken2', and their 120 recombinant inbred lines (RILs) under cold stress. A previously constructed high-density genetic linkage map was used for the mapping of the QTLs conferring cold tolerance at the plumule and seedling stages. A total of 4 QTLs for plumule-stage cold tolerance and 9 QTLs for seedling-stage cold tolerance were detected, with the maximum limit of detection reaching 5.20. Notably, a genetically overlapping QTL for both plumule and seedling stages was identified on chromosome 8, spanning a physical interval of 24 432 953–25 295 129 bp. Candidate genes within the detected QTL intervals were screened, and quantitative real-time polymerase chain reaction (qRT-PCR) was conducted to analyze the gene expression during the plumule and seedling stages. The results revealed that LOC_Os03g06570, LOC_Os03g07100, LOC_Os06g08280, LOC_Os08g38440, LOC_Os08g39100, and LOC_Os08g39540 exhibited significantly differential expression between the parental lines. These genes were either significantly downregulated or upregulated under cold stress. Among them, the first three gene (LOC_Os03g06570, LOC_Os03g07100, and LOC_Os06g08280) were hypothesized to be key candidates regulating the cold tolerance of rice seedlings, while the latter three genes (LOC_Os08g38440, LOC_Os08g39100, and LOC_Os08g39540) were identified as comprehensive regulators of cold tolerance during both plumule and seedling stages. These findings lay a foundation for the fine mapping and cloning of cold tolerance genes at the plumule and seedling stages, providing valuable insights for breeding cold-tolerant rice varieties.