大丽花花瓣衰老相关蛋白DpXTH1及其编码基因的分离

摘要

Objective] This study aimed to explore the molecular mechanism of senescence in ethylene-insensitive flowers. [Method] The dahlia petals were used as material, and the senescence-associated proteins were isolated and identified using two-dimensional electrophoresis, mass spectrometry and an encoding gene was cloned using molecular biology techniques. [Result] ln the two-dimensional elec-trophorogram of proteins from dahlia petals at building color, ful flowering and flow-er senescence periods, a total of 44 protein spots with differences in expression level more than two times were detected. From the 44 protein spots, xyloglucan endotransglycosylase/hydrolases (XTHs), a senescence-associated protein, was iso-lated and identified and its expression level was increased continuously with the senescence process of dahlia petals. By using the total RNA of dahlia petals as material and a pair of degenerate primers, the cDNA sequence of XTH gene was cloned by RT-PCR. The encoding region of XTH gene has a ful length of 882 bp, encoding 293 amino acid residues, and is named as DpXTH1 (Accession number:HM053613.1). The cluster analysis showed that the amino acid sequence of DpXTH1 has high homology with those of XTHs in other plants. [Conclusion] The isolated and identified DpXTH1 from dahlia petals belonged to the XTH family in plants, and its biological function was associated with the senescence process and regulation of dahlia petals.%[目的]为了探讨乙烯非敏感型花卉衰老相关的分子机理。[方法]以大丽花花瓣为材料，利用双向电泳、质谱分析及分子生物学技术，分离、鉴定花瓣衰老相关蛋白质及其编码基因。[结果]从大丽花透色期、盛花期和衰败期花瓣蛋白质的双向电泳图谱中，检测到44个表达量差异在2倍以上的蛋白质斑点，从中分离、鉴定了木葡聚糖内糖基转移酶/水解酶（xyloglucan endotransglycosylase/hydrolase, XTH），其表达量随花瓣的衰老进程而持续增加，属于一种衰老相关蛋白质。以大丽花花瓣RNA为材料，设计一对简并引物，通过 RT-PCR 技术克隆了大丽花花瓣 XTH 基因的cDNA序列，其编码区序列全长882 bp，编码293个氨基酸残基（基因登录号 HM053613.1，命名其为 DpXTH1）。聚类分析表明，DpXTH1氨基酸序列与其他植物的 XTH具有较高的同源性。[结论]分离、鉴定的大丽花 DpXTH1属于植物 XTH家族，其生物学功能与大丽花花瓣的衰老进程及调控有关。