家蚕DnaJ5基因的克隆与表达谱分析

Molecular Cloning and Expression Profile of Bm-DnaJ 5 Gene in the Silkworm,Bombyx mori

DnaJ蛋白是一种调节蛋白。从构建的家蚕蛹cDNA文库中检索到一条编码DnaJ蛋白的EST,并克隆获得家蚕DnaJ5基因(Bm-DnaJ5,GenBank登录号为FJ592078)。该基因的开放阅读框长1 056 bp,编码351个氨基酸,分子质量为38.93 kD,等电点为9.25。利用实时定量PCR技术对Bm-DnaJ5基因在家蚕不同组织和发育时期的时空表达谱进行定量分析,结果显示:该基因在5龄幼虫时期的各组织中都有表达,在睾丸中的表达量最高,拷贝数达到1.16×108个/μg,其次在头部、中肠、前部丝腺和翅原基中的表达量也较高,在气管丛中的表达量最低,拷贝数仅为1.84×104个/μg;在蛹中期的几个组织中均有表达,其中在眼和睾丸中的表达量较高,拷贝数分别为8.79×107个/μg和1.78×107个/μg,而在卵巢中的表达量最低,拷贝数仅为1.94×105个/μg。

DnaJ protein is a regulatory protein. A Bombyx mori EST was identified which encodes the DnaJ protein from silkworm pupa cDNA database and a cDNA sequence named Bm-DnaJ5 was cloned from the Bombyx mori. The deduced protein contains 351 amino acids, its molecular weight is 38.93 kD and theoretical pl is 9.25. The profile of spatio-temporal expression of Bm-DnaJ5 identified by real-time PCR indicated that it had a broad distribution in various tissues from the 5th instar larvae and pupae. Quantitative analysis showed that expression of Bm-DnaJ5 in the 5th instar larvae was the highest in testis （1. 16 × 10^8copy numbers/μg）, was relatively higher in brain, midgut, fore silk gland and wing discs, and was the lowest in tracheal cluster （1.84 ×10^4 copy numbers/μg）. The results also showed that Bm-DnaJ5 was expressed in six tissues of the middle pupal stage of Bombyx mori, with relatively higher levels （8.79 ×10^7copy numbers/pg and 1.78 ×10^7 copy numbers/μg） in eyes and testis and lowest level in ovary （1.94 × 10^5copy numbers/μg）.