发菜藻蓝蛋白基因克隆及原核表达

发菜（Nostoc flagelliforme）是一种陆生固氮蓝藻,具有重要的经济和生态价值。运用双向电泳技术、MALDI-TOF-TOF/MS鉴定和数据库检索,获得藻蓝蛋白部分氨基酸序列并设计简并性引物,克隆藻蓝蛋白基因并研究其表达。结果表明,发菜藻蓝蛋白α和β亚基两个基因的编码序列及两者之间的间隔序列全长为1097bp,编码β亚基和α亚基的基因序列全长分别为519bp和489bp,β亚基基因序列位于α亚基基因序列上游,两者之间通过89bp的基因片段连接,GenBank登录号为GU549478,并对推译的α和β亚基三维结构进行了预测。将藻蓝蛋白的α和β亚基基因在大肠杆菌中表达,获得了符合预期的外源重组蛋白。研究结果为进一步研究发菜藻蓝蛋白的分子结构及生物学功能奠定了基础。

Cloning and Prokaryotic Expression of C-Phycocyanin Gene from Nostoc flagelliforme

Nostoc flagelliforme is a kind of terrestrial nitrogen-fixing cyanobacteria with important economic and ecological value.2-DE method,gel image analysis and MALDI-TOF-TOF/MS were used to identify C-Phycocyanin.Gene was cloned with designed degeneracy primer by known amino acid sequences.The results indicated that the full length of C-Phycocyanin was with 1097 bp（GenBank access number GU 549478）.The subunit β gene-coding region contains 519 bp and the subunit α gene-coding region contains 489 bp with a 89 bp spacer between them.The 3D structure model of subunit α and β of N.flagelliforme were prognosticated.Subunit α and β of C-Phycocyanin gene was expressed in E.coli,and heterologous proteins were obtained.The results laid a foundation for further study on molecular structure and biological functions of C-Phycocyanin from N.flagelliform.