四川小麦地方品种AS1643中α/β-醇溶蛋白基因的序列分析

Sequence Analysis of α/β-gliadin Genes from Sichuan Wheat Landrace AS1643

【目的】克隆四川小麦地方品种AS1643中的α/β-醇溶蛋白基因,并进行序列分析,探讨小麦AS1643的优良品质与其贮藏蛋白基因间的关系。【方法】采用PCR扩增的方法。【结果】从四川小麦地方品种AS1643中克隆到3个α/β-醇溶蛋白基因,即Gli-AS1643-1(GenBankNo.DQ166376)、Gli-AS1643-2(GenBankNo.DQ166377)和Gli-AS1643-3(GenBankNo.DQ166378)。其中,Gli-AS1643-1和Gli-AS1643-2的编码区长度分别为873和852bp,可编码270和263个氨基酸残基的成熟蛋白。Gli-AS1643-3由于在编码区的第535～537位存在一个提前终止密码子,推测为不能编码成熟蛋白的假基因。序列比较显示Gli-AS1643-1、Gli-AS1643-2和Gli-AS1643-3分别与GenBank中的α/β-醇溶蛋白基因具有较高的一致性,且序列结构非常相似。它们的N-端氨基酸序列与各种α-、β-、γ-和α/β-醇溶蛋白的基本一致,但与ω-醇溶蛋白和低分子量谷蛋白亚基的明显不同。在N-端具有12肽串联重复序列特征,即紧密相关的5个脯氨酸框,在Ⅱ区和Ⅳ区具有类似于微卫星序列编码的2个多聚谷氨酰胺区域。在Gli-AS1643-2N-端存在与腹泻疾病相关的序列,C-端含有12型腺病毒感染序列。Gli-AS1643-1、Gli-AS1643-2和Gli-AS1643-3中都有可形成3个分子内二硫键的6个保守的半胱氨酸残基。【结论】小麦AS1643具有较好品质可能在于其贮藏蛋白基因间的相互作用等所致。

[Objective] The objective was to clone α/β-gliadin genes from Sichuan wheat landrace AS1643, to analyze their gene sequences and to explore relationship between good quality of wheat AS1643 and its protein-storing gene. [Method] Method of PCR amplification was used in this study. [Result] The full coding regions （open reading frame, ORF） of three α/β-gliadin genes Gli-AS1643-1 （GenBank No.DQ166376）, Gli-AS1643-2 （GenBank No.DQ166377） and GIi-AS1643-3 （GenBank No. DQ166378）, were isolated from the genomic DNA of Sichuan wheat landrace AS1643. Among which, Gli-AS1643-1 and Gli-AS1643-2 were 873 bp and 852 bp, and could encode two mature proteins with 270 and 263 amino acid residues, respectively. Due to one stop codon in the 535-537 positions of its coding region, Gli-AS1643-3 was speculated to be a pseudogene. Multiple sequence alignment analysis suggested that Gli-AS1643-1, Gli-AS1643-2 and Gli-AS1643-3 have the higher similarity in their structure with the known α/β-gliadin genes in GenBank. N-terminal amino acid sequences of the three cloned genes were basically consistent with other α-, β-, γ- and α/β-gliadins, whereas it was clearly different with ω-gliadins and LMW-GS. N-terminal amino acid sequence was dodecapeptide tandem repeat feature with five more closely proline boxes. In Ⅱ and Ⅳ regions were two polyglutamine domains encoded by microsatellite-like sequences. Sequence active in celiac disease and adenovirus type twelve infection sequences were presented in N-domain and in C-domain of Gli-AS1643-2, respectively. The six conserved cysteine residues could form three intramolecular disulfide bonds in Gli-AS1643-1, Gli-AS1643-2 and Gli-AS1643-3, respectively. [ Conclusion ] Good quality of wheat AS 1643 could result form mutual effect of their storage protein or other factors.