家蚕细胞周期蛋白基因BmCcnl1的克隆及表达分析  

Cloning and expression analysis of the cyclin gene BmCcnl1 in the silkworm,Bombyx mori

作　　者：范兰芬[1] 钟杨生[1] 林健荣[1] 

机构地区：[1]华南农业大学动物科学学院,广州510642

出　　处：《昆虫学报》2010年第12期1325-1332,共8页Acta Entomologica Sinica

基　　金：国家自然科学基金项目(30771634)

摘　　要：家蚕Bombyxmori由受精卵到完成胚胎发育孵化的过程中,细胞进行大量的分裂和分化,然而滞育性卵的胚胎细胞分化至G2期便停滞在此阶段。为了探索这一发育阶段细胞内的分子调控,本研究以人Homo sapiens的细胞周期蛋白基因cyclin L1为模板,成功克隆了家蚕同源基因BmCcnl1(GenBank登录号:FJ889988)。BmCcnl1基因开放阅读框(open reading frame,ORF)全长1254bp,编码417个氨基酸。利用Protean软件分析得出BmCcnl1蛋白预测分子量为49kDa,等电点为9.84。利用DNA重组技术构建了BmCcnl1基因的重组表达载体pET-21d-BmCcnl1,对其进行原核表达,其表达的蛋白以包涵体形式存在。利用RT-PCR技术分析了BmCcnl1基因在胚胎发育过程中的转录水平,BmCcnl1基因在非滞育性卵的胚胎发育阶段基本保持相对稳定的转录表达,而滞育性卵从蛾体产下经过72h后已经检测不到BmCcnl1基因的转录。结果提示,BmCcnl1基因与胚胎期滞育及非滞育性卵的发育调控相关。对该基因的克隆和表达分析为今后研究家蚕胚胎发育及细胞周期调控奠定了基础。Cell division and differentiation were processing in quantity from the fertilized eggs to larvae of the silkworm,Bombyx mori,but in diapause eggs embryonic development arrested in the G2 of cell division. In order to explore the molecular regulation in this developmental stage,the cyclin gene of B. mori was cloned according to the cyclin L1 sequence of Homo sapiens and named BmCcnl1. The ORF of BmCcnl1 is 1 254 bp,encoding a 417-amino acid protein and registered with GenBank accession number FJ889988. The predicted molecular weight and isoelectric point of BmCcnl1 protein were 49 kDa and 9.84,respectively analyzed by Protean software. The prokaryotic expression vector pET-21d-BmCcnl1 was constructed,and the expression product was present in inclusion bodies. The expression of the BmCcnl1 mRNA was detected using RT-PCR. The relative expression level of BmCcnl1 mRNA was stable in the embryonic stage of non-diapause eggs,but its expression was not detectable 72 h after oviposition. The results suggest that BmCcnl1 is related to the development regulation of the diapause and non-diapause eggs. The cloning and expression analysis of BmCcnl1 provide the basis for the research of the embryonic development and cell cycle regulation.

关 键 词：家蚕 细胞周期蛋白 克隆 表达 滞育 

分 类 号：Q965[生物学—昆虫学]