东方山羊豆水通道蛋白基因的克隆及初步分析

摘要

Based on a deposited EST fragment, the full-length cDNA of one novel gene encoding aquaporin had been cloned from Galega Orientalis using RT-PCR and RACE technology. The new gene was identified as the GoAQP gene and had been assigned GenBank accession number: HM803185. Bioinformatics analysis showed that the full- length cDNA of this gene is 1258 bp including an 870-bp open reading frame (ORF), which encodes a protein of 289 amino acids. GoAQP protein possesses the conserved MIP family signal sequence HINPAVT/SFG, the highly conserved sequence of the higher plant GGANXXXXGY and TGI/TNPARSL/FGAAI/VI/VF/YN. Compared to the AQP family from Arabidopsis thaliana, phylogenetic analysis indicated that GoAQP belong to PIP1 subfamily. Real-Time PCR results revealed that expression of GoAQP was the most abundant in root and the least in stem. The expression level of GoAQP in leaves were increased with NaCI and PEG stress, and showed a bimodal distribution pattern in NaC1 treatment. This suggested that GoAQP might be involved in the manipulation of salt tolerance of Galega Orientalis. The expression plasmid pCAMBIA-i302-GoAQP was constructed for the further gene function analysis using the transgenic technology.%根据已知EST片段结合RT-PCR和RACE技术,从东方山羊豆(Galega Orientalis)中克隆到一个水通道蛋白(AQP)基因,命名为GoAQP(GenBank登录号:HM 803185).测序和生物信息学分析表明,此序列全长1258bp,开放读码框(ORF)870 bp,编码289个氨基酸.GoAQP包含MIP家族信号序列HINPAVT/SFG,高等植物PIP高度保守序列GGANXXXXGY和TGI/TNPARSL/FGAAI/VI/VF/YN.与拟南芥AQP基因家族的系统进化分析表明,GoAQP属于PIP1亚型AQP蛋白.Real-Time PCR检测结果表明,GoAQP基冈在根中表达量最高,茎中表达量最少;且受NaCl和PEG胁迫表达上调,NaCl表达呈双峰分布.因此GoAQP在基因可能参与了东方山羊豆耐盐性调控.同时成功构建超表达载体pCAMBIA-1302-GDAQP,为转基因验证基因功能创造了基础.