普通小麦及近缘粗山羊草α-醇溶蛋白基因的克隆、定位与进化分析

Cloning,Chromosomal Location,and Evolutionary Analysis of α-gliadin Genes from Aegilops tauschii and Common Wheat (Triticum aestivum L.)

通过特异PCR引物设计,从普通小麦品种(豫麦34和烟农19)和粗山羊草(T9、T197、T48、T176和T17)中扩增、克隆了7个新的α-醇溶蛋白基因,分别命名为Gli-YM34、Gli-YN19、Gli-T9、Gli-T197、Gli-T48、Gli-T176和Gli-T17,基因序列长度为846～891bp,编码282～297个氨基酸残基,都具有α-醇溶蛋白的典型结构特点。其中Gli-YM34和Gli-YN19基因推导的醇溶蛋白都含有一个额外的半胱氨酸残基,可能对面筋品质有正向作用。根据α-醇溶蛋白氨基酸序列所具有的4种T细胞抗原表位和多聚谷氨酰胺重复区的平均长度以及中国春缺体四体分析,将来自普通小麦品种的Gli-YM34和Gli-YN19基因定位在6D染色体上的Gli-D2位点,而且Gli-YM34和Gli-YN19与来自粗山羊草的α-醇溶蛋白基因具有很高的序列相似性,进一步证明粗山羊草是普通小麦D基因组的供体。在克隆的4个典型α-醇溶蛋白基因中检测到21个SNP和1个9bp的缺失。系统进化分析表明,α-醇溶蛋白基因与低分子量谷蛋白亚基基因关系较近,在大约43.69百万年时分化,与ω-醇溶蛋白和HMW-GS基因亲缘关系较远,它们的分化时间大约为79.39百万年。

Seven novel α-gliadin genes from common wheat cultivars （Yumai 34 and Yannong 19） and Aegilops tauschii accessions （T9,T197,T48,T176,and T17） were amplified and cloned by using a PCR-based strategy. They were designated as Gli-YM34,Gli-YN19,Gli-T9,Gli-T197,Gli-T48,Gli-T176,and Gli-T17,respectively. Their length of the open reading frame （ORF） ranged from 846 to 891 bp,encoding the putative proteins of 282–297 amino acid residues. Comparative analysis showed that all genes isolated had typical structural characters of α-gliadin genes reported previously. Particularly,Gli-YM34 and Gli-YN19 α-gliadins from common wheat possessed an additional cysteine residue,suggesting a possible positive effect on dough quality. Both genes were assigned to Gli-D2 locus on the chromosome 6D by the analysis of four celiac disease toxic epitopes and glutamine residues in the polyglutamine domain as well as nullisomic-tetrasomic lines of Chinese Spring. A total of twenty-one single nucleotide polymorphisms （SNPs） and a 9 bp deletion among the four typical α-gliadin genes were identified. Phylogenetic and evolutionary analysis revealed that the α-gliadin genes are closely related to LMW-GS genes and their divergence occurred 43.69 million year ago. Less homology was found between α-gliadin genes and HMW-GS and ω-gliadin genes,and they diverged about 79.39 million year.