At3g13600截短型蛋白的原核表达载体的构建

Construction of the recombinant prokaryotic expression plasmids contain truncated form of At3g13600 protein

拟南芥基因At3g13600编码一种钙调素结合蛋白,序列分析表明,该基因cDNA序列全长1 818 bp,编码一个具有606个氨基酸残基的多肽,推测分子量为68.9 kD;在At3g13600蛋白的N端存在IQ钙调素结合构象.为了从实验上进一步研究该蛋白的钙调素结合特性,通过逆转录聚合酶链式反应(RT-PCR)扩增含IQ序列与否的不同长度的编码区cDNA序列,构建到原核表达载体pET32 a+中.测序结果表明:目的序列已正确克隆到表达载体上,为进一步表达目的蛋白及研究其功能奠定了基础.

Arabidopsis gene At3g13600 encoded a calmodulin-binding protein. Sequence analysis showed that the full length cDNA was 1 818 bp and encoded a polypeptide containing 606 amino acid residues,its molecular weight is about 68.9 kD. Amino acid sequence of the deduced protein revealed the existence of IQ calmodulinbinding motif at the N-terminus. To further study the calmodulin binding characteristics of tile protein, the coding domain of different length eDNAs were reverse amplified by polymerse chain reaction （PCR） , and subcloned into prokaryotic expression vector pET32a ＋. DNA sequencing showed that the different length cDNAs had been correctly inserted into the reeombinants plasmids. This experiment will build up a foundation for further studying the expression of the target protein and identification of its functions.