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根据生物信息学分析,同源性克隆分离玉米DCL1候选基因,鉴定该基因组织与逆境响应等表达特征,并分析其自然等位变异。结果表明,目标候选基因全长7 408 bp,与拟南芥和水稻DCL1基因氨基酸序列高度保守,相似度分别为91%和77%。mRNA定量PCR分析表明,该基因在幼叶和吐丝期雌穗中表达丰度分别是幼茎的75.35倍和16.21倍;高盐、脱水、ABA、低氮、低磷胁迫处理条件下,幼苗中高盐和ABA处理呈下调表达,低氮和低磷处理呈显著上调表达,脱水处理无显著变化。核苷酸与氨基酸序列的保守性、组织与逆境响应差异表达特征均与拟南芥和水稻等相似。87份玉米自交系10×全基因重测序数据分析结果表明,该基因遗传变异丰富,共83个位点存在自然变异,但外显子区域和主要的功能结构域序列相对保守,约2/3的自然变异集中在内含子区域,外显子区域的27个自然变异位点只有8个位点在16个材料存在氨基酸差异。
According to the bioinformatics analysis, homologous clones were used to isolate the candidate gene of maize DCL1, identify the expression characteristics of the gene and its response to stress, and analyze its natural allelic variation. The results showed that the target gene was 7 408 bp in length, which was highly conserved among the amino acid sequences of Arabidopsis and rice DCL1 gene. The similarity was 91% and 77% respectively. Quantitative PCR analysis showed that the expression level of this gene was 75.35 times and 16.21 times higher in young ear and silking ear, respectively. Under high salt, dehydration, ABA, low nitrogen and low phosphorus stress, High and low salt and ABA treatments were down-regulated, low and low-phosphorus treatments were up-regulated and dehydration treatments did not change significantly. The conservation of nucleotide and amino acid sequence, the differential expression between tissue and adversity responses are similar to that of Arabidopsis and rice. The analysis of 10 × whole genome sequencing data in 87 maize inbred lines showed that there are abundant genetic variations in 83 maize inbred lines, of which 83 loci have natural variation. However, the exon region and the major functional domain sequences are relatively conservative, about 2 / 3 of the natural variation concentrated in the intron region, exon region 27 natural variation sites only 8 sites in 16 material amino acid differences exist.