拟南芥转录因子CBF1基因的克隆与原核表达

Cloning and Prokaryotic Expression of Transcription Factor CBF1 Gene in Arabidopsis thaliana

CBF1转录激活因子与植物的抗逆密切相关,根据GenBank中拟南芥(Arabidopsis thaliana)CBF1(C-repeat/dre binding factor 1)基因的编码区设计1对特异引物,通过RT-PCR扩增出拟南芥CBF1基因,随后亚克隆到pMD18-T载体,测序、序列分析表明与拟南芥CBF1对应区域相似性为100%。将该片段亚克隆到原核表达载体pET22b(+)中构建原核表达质粒,转化大肠杆菌ER2566,在IPTG诱导下产生CBF1蛋白,SDS-PAGE结果表明表达的蛋白约24ku,与预期一致。

CBF1 transcription factor is very important for plants to bear adversity. A pair of PCR primers was designed and synthesized according to the sequences of CBF1 gene of Arabidopsis thaliana published in GenBank. The ORF of CBF1 was amplified with RT-PCR. After ligation,transformation and sequence,the segment was identical with the CBF1 of A. thaliana in GenBank. Then CBF1 segment was subcloned into pET22b(+) and an expressional plasmid of pET22b-CBF1 was constructed.The plasmid was transformed into E.coli ER2566,and was induced with IPTG.SDS-PAGE result showed that a light strip about 24 ku was expressed,which was predicated.