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【目的】探索水稻剑叶叶宽调控及其对氮肥响应的遗传机理,为氮高效水稻新品种的培育提供新的种质资源和基因标记。【方法】以134份水稻地方种质资源为关联分析材料,通过基因组重测序发掘获得了3 356 591个分布于全基因组的高密度SNP位点(single nucleotide polymorphism,SNP)。在大田栽培条件下,以施氮量为主区,品种为裂区的设计,设计低氮(不施氮肥,N0)、正常氮(纯氮96 kg·hm~(-2),N1)和高氮(纯氮192 kg·hm~(-2),N2)3种氮肥处理。于水稻成熟期分别调查水稻剑叶叶宽在低、中、高3种氮肥处理下的表现及响应,结合EMMAX软件计算群体亲缘关系矩阵和EIGENSOFT软件分析群体结构,采用纳入亲缘关系矩阵及群体结构的混合线性模型开展全基因组关联分析。【结果】水稻剑叶叶宽在N0、N1、N2 3种氮肥处理下均呈正态分布,并表现丰富的变异。剑叶叶宽受品种差异及氮水平的影响,且与施氮水平呈显著正相关。在N0、N1、N2氮处理下共检测到14个与剑叶叶宽显著相关的SNP位点。其中,低氮处理下检测到的SNP位点的最小等位基因频率均大于0.46,表明此类SNP在关联群体中广泛存在;中氮和高氮水平下检测到的SNP位点的最小等位基因频率均较小,是一类较为稀有的SNP位点。位于第12染色体上的一个SNP(chr12:15 066 507)位点在正常氮及高氮处理下均被检测到,在高氮处理下还检测到的另一显著性位点,其候选区间内包含一候选基因LOC_Os12g25660,该基因与业已报道的叶宽性状相关基因Os BR6ox同属于细胞色素P450家族。根据不同氮处理下剑叶叶宽的响应,鉴定出20与低氮响应有关的SNP位点,8个位点与高氮响应有关。其中与高氮响应的显著性位点中,位于第1染色体显著性峰候选区间包含业已克隆的与氮素利用相关的基因OsATG7。【结论】通过全基因组关联分析共检测到42个与剑叶叶宽及其在不同氮处理下叶宽响应相关的显著性关联位点。
【Objective】 The aim of the study was to explore the genetic regulation of flag leaf width and its response to nitrogen fertilizer in rice and to provide new germplasm resources and genetic markers for the cultivation of new high efficient nitrogenous rice varieties. 【Method】 With the analysis of 134 rice germplasm resources, 3 356 591 high-density SNP loci distributed in the whole genome were discovered through genome resequencing. Under the condition of field cultivation, nitrogen (nitrogenous fertilizer, N0), normal nitrogen (pure nitrogen 96 kg · hm -2, N1) and High nitrogen (pure nitrogen 192 kg · hm -2, N2) three nitrogen treatments. At the rice ripening stage, the performance and response of rice flag leaf width under low, medium and high nitrogen fertilizer treatments were investigated respectively. Combined with EMMAX software to calculate population genetic relationship matrix and EIGENSOFT software to analyze population structure, genetic relationship and population structure A hybrid linear model for genome-wide association analysis. 【Result】 The leaf width of flag leaf in rice was normal distribution under N0, N1 and N2 treatments and showed abundant variation. The width of flag leaf was affected by the difference of cultivars and nitrogen level, and was positively correlated with the nitrogen level. A total of 14 SNPs significantly associated with flag leaf width were detected under nitrogen, nitrogen and nitrogen treatments. Among them, the minimum allele frequency of SNP loci detected under low nitrogen treatment was more than 0.46, indicating that such SNPs were widely found in related populations; the lowest allele of SNP loci detected under medium nitrogen and high nitrogen levels Gene frequency are small, is a relatively rare SNP sites. One SNP (chr12: 15 066 507) locus on chromosome 12 was detected under both normal nitrogen and high nitrogen treatments. Another significant site detected under high nitrogen treatment was within the candidate interval It contains a candidate gene LOC_Os12g25660, which belongs to the cytochrome P450 family and has been reported to belong to the cytochrome P450 family. According to the responses of flag leaf width under different nitrogen treatments, 20 SNPs related to low nitrogen response were identified, of which 8 were related to high nitrogen response. Among the significant sites responding to high nitrogen, the candidate region on the chromosome 1 significant peak contains the cloned gene of nitrogen utilization related OsATG7. 【Conclusion】 A total of 42 significant correlation sites were detected by genome-wide association analysis, which were related to leaf width of flag leaf and leaf width response under different nitrogen treatments.