斑节对虾细胞周期蛋白T基因的克隆和表达分析

Molecular cloning and expression of cyclin T gene from black tiger shrimp( Penaeus monodon)

真核生物的转录调控多发生在转录的延伸阶段,细胞周期蛋白T(cyclin T)是细胞转录的重要调控因子。文章利用RACE技术获得斑节对虾(Penaeus monodon)cyclin T(Pmcyclin T)c DNA全长,其全长为3 421 bp,其中开放阅读框(ORF)3 135 bp,编码1 044个氨基酸;生物信息学分析显示,该氨基酸序列含有一个周期蛋白家族特有的CYCLIN保守区,并含有N-糖基化位点和磷酸化位点;经BLAST同源性分析显示,细胞周期蛋白T编码的蛋白与柑橘木虱(Diaphorina citri)、切叶蚁(Acromyrmex echinatior)等多种节肢动物有很高的同源性;利用qRTPCR技术对Pmcyclin T在不同组织及卵巢发育各时期的mRNA水平的表达谱进行了研究。结果表明,细胞周期蛋白T在斑节对虾的心、卵巢等7个组织中均有表达,其中卵巢中表达量显著高于其他组织(P<0.05);在卵巢发育各时期中,细胞周期蛋白T在Ⅲ期表达量最高。

Cyclin T is a critical factor that stimulates the process of transcription elongation. We obtained the full-length c DNA sequence of cyclin T from black tiger shrimps（ Penaeus monodon）（ denoted as Pmcyclin T） by RACE PCR. The c DNA of Pmcyclin T was 3 421 bp,encoding a polypeptide of 1 044 amino acids. Bioinformatics analysis shows that the amino acid sequence contained a CYCLIN conserved region of a periodic protein family as well as N-glycosylation sites and phosphorylation sites. The results of Blast and phylogenetic analyses suggest that Pmcyclin T shared very high homology with other kinds of arthropod such asDiaphorina citri,Acromyrmex echinatior and so on. Besides,we measured the mRNA expression of Pmcyclin T in seven tissues by real-time PCR. Expression was highest in the ovary and moderate in the intestine,brain and so on. While at different stages of ovary,Pmcyclin T was highest expressed at Stage Ⅲ,suggesting that Pmcyclin T might play an important role in ovary development of black tiger shrimp.