毛竹PsbS基因的克隆及其原核表达特征研究

Cloning of PsbS Gene from Moso Bamboo (Phyllostachys edulis) and Its Characterization of Prokaryotic Expression

摘要:

采用RT-PCR技术从毛竹(Phyllostachys edulis)叶片中克隆到1个PsbS基因,命名为PePsbS (GenBank No. FJ600727),其编码区为810 bp,编码269个氨基酸。序列分析表明,PePsbS编码的蛋白与其它单子叶植物的PsbS蛋白有很高的相似性。蛋白结构分析表明,PePsbS基因编码蛋白包含导肽部分和成熟蛋白,其中成熟蛋白包含4个跨膜结构域。将PePsbS基因编码成熟蛋白的序列构建到原核表达载体pET23a中,并转入大肠杆菌,用IPTG进行诱导表达。结果表明, 41℃下诱导4 h的表达效果最好,目的蛋白含量约占总蛋白的21.5%,分子量约为22.0 kD。这说明温度和诱导时间明显影响PePsbS基因的表达。

Abstract:

PsbS protein plays an important role in non-photochemical quenching (NPQ) in plants. A PsbS gene was cloned from Moso bamboo (Phyllostachys edulis) leaves by RT-PCR method, which named PePsbS (GenBank No. FJ600727) with 810 bp in length, and encoded a polypeptide with 269 amino acids. Homology analysis showed that the deduced PsbS was highly homologous to those from other monocotyledon plants. The protein structure analysis indicated that PePsbS consists of transit peptide and mature protein including four transmembrane domains. The fragment of PePsbS gene encoding mature protein was constructed in multiple cloning site of pET23a and expressed in Escherichia coli induced by IPTG. The expression of PePsbS was the best under 41℃ for 4 hours, the recombinant protein accounted for 21.5% of the total proteins with molecular weight about 22 kD. It indicated that there were significant effects of temperature and IPTG induced time on protein expression.