文章摘要
杨亚男,黄辉洋,黄小帅,叶海辉,李少菁.近缘新对虾cyclin B基因的克隆与原核表达[J].水产学报,2013,37(2):184~191
近缘新对虾cyclin B基因的克隆与原核表达
Molecular cloning and prokaryotic expression of cyclin B from shrimp Metapenaeus affinis
投稿时间:2012-06-14  修订日期:2012-11-23
DOI:DOI:10.3724/SP.J.1231.2013.38210
中文关键词: 近缘新对虾  cyclin B基因  克隆  原核表达
英文关键词: Metapenaeus affinis  cyclin B  gene cloning  prokaryotic expression
基金项目:国家自然科学基金项目(40406030)
作者单位
杨亚男 厦门大学海洋与地球学院 
黄辉洋 厦门大学海洋与地球学院 
黄小帅 厦门大学海洋与地球学院 
叶海辉 厦门大学海洋与地球学院 
李少菁 厦门大学海洋与地球学院 
摘要点击次数: 2353
全文下载次数: 1263
中文摘要:
      采用RACE技术,克隆了近缘新对虾cyclin B基因,该序列全长1 667 bp,编码区1 209 bp共编码402个氨基酸,所推导的氨基酸序列N端不存在信号肽。BLAST比对后发现,其氨基酸序列与刀额新对虾的同源性达90%。经RT-PCR检测,cyclin B基因在近缘新对虾的卵巢和肌肉中表达水平最高,胸神经节和心脏次之,鳃、眼柄神经节、肝胰腺几乎没有表达。推测主要与cyclin B在细胞周期中调控细胞分裂的功能有关。原核表达结果显示,pET32a表达载体在0.2 mmol/L IPTG、25 ℃诱导4 h条件下可得到纯度较高的分子量约67 ku的蛋白。
英文摘要:
      Cyclin B gene was cloned from shrimp Metapenaeus affinis using RACE.The results showed that the full-length cDNA of cyclin B gene is 1 667 bp,including 1 209 bp coding sequence,the open reading frame encodes a putative peptide of 402 amino acid.The encoded protein does not contain signal peptides in this sequence.BLAST search has shown that the deduced peptide shared 90% sequence identity with that of Metapenaeus ensis.Gene expression profiling by semi quantitative reverse transcriptase polymerase chain reaction(RT- PCR)demonstrated that cyclin B expression levels are the highest in ovary and muscle,then in thoracic ganglion and heart,but almost no expression in gill,hepatopancreas and eyestalk.The results suggest that cyclin B expression levels may relate to its function as a regulator of cell division during cell cycle.Result of prokaryotic expression showed that,the recombinant protein was expressed with expression plasmid pET32a.At last we got a high quantity of recombinant protein(67 ku)in the optimum induction conditions,when the concentration of IPTG was 0.2 mmol/L for 4 h at 25 ℃ for 32a-MaCB.
HTML   查看全文   下载PDF阅读器
关闭

手机扫一扫看