齿肋赤藓乙醛脱氢酶基因ALDH21的克隆与表达分析

本文采用RT-PCR方法,从齿肋赤藓(Syntrichia caninervis)总RNA中获得ALDH21基因cDNA序列,连接到pMD19-T载体上并转化E.coli DH5α,阳性克隆经PCR鉴定后测序,将测序结果与GenBank中山墙藓(Tortula ruralis)和小立碗藓(Physcomitrella patens)相关基因序列进行同源性比对。结果表明,实验成功克隆了S.caninervis的ALDH21基因的cDNA序列(GenBank登录号:GQ245973),为一个完整的ORF,其长度为1452bp。该序列与T.ruralis和P.patens的cDNA序列同源性分别为98%和78%,推导的氨基酸同源性分别为97%和87%。生物信息学分析表明该蛋白质分子量为52.98kD,等电点pI为5.96,编码483个氨基酸,具备ALDH21蛋白家族特征;半定量RT-PCR结果表明:ALDH21在干旱胁迫状态时的表达量显著高于水合状态,说明ALDH21基因可能参与干旱胁迫应答。本文为进一步研究齿肋赤藓ALDH21基因的抗旱机理提供理论依据。

Cloning of ALDH21 from Syntrichia caninervis and Expressing Analysis

In this research, we obtained the ALDH21 gene cDNA sequence from the total RNA of Syntrichia ca-ninervis with the RT-PCR method, and then was ligased into the pMD19-T vector, and was transformed into E. coli DH5α.After identified by PCR, the positive clones were sequenced. Sequencing results were compared with the correlation gene sequence of Tortula rura//s and Physcomitrella patens in GenBank. The result demonstrates that we have successfully cloned an ALDH21 gene cDNA sequence from S. eaninervis, with the GenBank accession nummber is GQ245973, which containns the gene ORF with 1452 bp. The nucleotide sequence homology com-pared to the cDNA sequence of T. ruralis and P. patens is 98% and 78%, respectively, and the deduced amino acid homology is 97% and 87%, respectively. Bioinformation analysis displays that the protein molecular weight is 52.98 kD and the isoelectric point pI is 5.96, encodes 483 amino acids and belongs to ALDH21 protein family.The result of semi-quantitative RT-PCR shows that the expression ofA LDH21 gene when S. caninervis is desicca-tion is significantly higher than that of when is hydration, which illustrates that the ALDH21 gene might involve in the desiccation-tolerant responsion. This paper would lay the foundation for further studying the desiccation-toler-ant mechanism of the A LDH21 gene in Syntrichia caninervis.