高粱LEA3蛋白基因和启动子的克隆及序列分析

doi:10.7677/ynzwyj201312110

摘要/Abstract

摘要：

根据禾本科LEA3基因保守序列设计简并引物，同时结合RACE方法获得高粱LEA3基因全长cDNA序列1032bp，该序列含有一个612bp的阅读框，编码203个氨基酸，包含7个串联的LEA3蛋白的基元序列。通过与玉米、小麦、水稻、大麦的LEA3蛋白序列比较，氨基酸序列同源性分别为73.8%、53.77%、45.63%和53.99%；其编码蛋白理论相对分子量为21.22kD，等电点pI=8.79；蛋白质二级结构预测表明2段α螺旋结构占主导，与目前已知的多种植物的LEA3蛋白具有相似的结构功能域。通过热不对称交错PCR(TAILPCR)技术获得LEA3基因启动子749bp的DNA序列，该区域包含ABA应答元件、干旱胁迫应答元件、以及胚胎和胚乳特异表达元件；通过PHyML软件构建了禾本科植物LEA3基因ML系统树。这些研究结果为深入了解该基因的功能和高粱抗旱的分子机理提供了基础数据。

关键词: 高粱, LEA3蛋白, 克隆, 启动子, ML树

Abstract:

A pair of degenerate primers was designed according to reported fragments of LEA3 genes in GenBank. Using these primers a new cDNA fragment of 1032bp was amplified by PCR and RACE from total RNA of Sorghum bicolor, which indicated that the open reading frame region of the gene was 612bp (GenBank accession number GQ494000). It is predicted that the sequence encodes 203 amino acids with a molecular weight of about 21.22kD, includes seven characteristic motifs of LEA3 and has a theoretical isoelectric point of 8.79. The sequence shows a similarity of 73.8%, 53.77%, 45.63% and 53.99% with Zea mays, Triticum aestivum, Oryza sativa and Hordeum vulgare LEA3 sequences, respectively. The secondary structure of the Sorghum LEA3 protein exhibited a quantity of αhelix dominant structures that were similar to those found in many other plants. In addition, a 749bp DNA sequence was isolated from Sorghum genomic DNA by Thermal Asymmetric Interlaced PCR (TailPCR). This promoter region was predicted to contain ABA and drought stress response elements of embryo and endospermspecific genes Sequences of Gramineae LEA3 genes were aligned using the profile alignment function of ClustalX, and maximum likelihood phylogenetic analyses were conducted using PHyML. The research forms part of an analysis of droughtresistant mechanisms and gene function in Sorghum.

Key words: Sorghum bicolor, LEA3 protein, Clone, Promoter, ML tree

中图分类号:

Q 78

孙晓娇, 汤晓倩, 于丽霞, 白琼, 鄢波. 高粱LEA3蛋白基因和启动子的克隆及序列分析[J]. Plant Diversity, 2013, 35(5): 585-593.

SUN Xiao-Jiao, SHANG Xiao-Qian, XU Li-Xia, BAI Qiong, YAN Bo. Cloning and Characterization of the LEA3 Protein Gene and Promoter from Sorghum bicolor[J]. Plant Diversity, 2013, 35(5): 585-593.

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