BmLSP基因启动子驱动DsRed在转基因家蚕中的表达分析

家蚕幼虫血清蛋白(BmLSP)是由脂肪体细胞合成的一种贮藏蛋白,是研究家蚕幼虫期发育调控的理想靶标。基于家蚕基因组数据,克隆了BmLSP基因5'端侧翼区～1.6 kb的调控序列,构建成以红色荧光蛋白基因DsRed为报告基因的pig-gyBac表达载体并进行转基因注射,在个体水平上验证该启动子的特性。结果表明:BmLSP-DsRed表达框以单拷贝形式整合至家蚕基因组;DsRed在转基因家蚕脂肪体中特异转录表达,其时期表达特征与BmLSP基因基本一致,随发育阶段的不同呈现有规律的变化,即幼虫期表达,但1龄、2龄眠期不表达,3龄、4龄眠期有微弱表达,上蔟后表达量逐渐降低直至化蛾阶段消失。提示BmLSP可能通过其启动子区的特异元件受激素的精确调控而行使功能。

Expression Analysis of DsRed Gene Driven by Promoter of BmLSP Gene in Transgenic Silkworm,Bombyx mori

The storage protein,Bombyx mori larval serum protein(BmLSP),is synthesized in the fat body of silkworm and is considered one of the target molecules for studying the regulation of larval development.Based on the data of silkworm genome,a 1.6 kb promoter sequence was cloned from the 5′ flanking region of BmLSP gene to construct a piggyBac expression vector by using DsRed as a reporter gene,which was further used to conduct transgenic microinjection for verification of the promoter's characters in individual silkworm larvae.The results showed that the reading frame of BmLSP-DsRed was inserted into the genome of silkworm in a single copy form and was expressed in a fat body-specific pattern in transgenic individuals.Moreover,its temporal expression pattern was similar to that of the endogenous BmLSP gene and exhibited a regular variation in different stages of development.To be specifically,high expression was observed during the larval stage,no expression was detected in molting larvae of the 1st and 2nd instars,weak expression was observed in molting larvae of the 3rd and 4th instars,the expressed product was reduced gradually after spinning and disappeared at the eclosion stage.These results suggest that the expression of BmLSP gene might be regulated accurately by some factors such as juvenile hormone that bind to specific elements of the promoter.