家蚕滞育激素基因的克隆及在多化性和二化性品种间的表达差异  

Molecular Cloning and Differential Expression of Diapause Horm one Gene Between Multivoltine and Bivoltine Silkworm Varieties

作　　者：何庆玲[1,2] 钱荷英[2] 许平震[1,2] 孙平江[2] 李刚[2] 张月华[2] 徐安英[2] 

机构地区：[1]江苏科技大学生物技术学院,江苏镇江212018 [2]中国农业科学院蚕业研究所,江苏镇江212018

出　　处：《蚕业科学》2015年第3期437-441,共5页ACTA SERICOLOGICA SINICA

基　　金：现代农业产业技术体系建设专项(No.CARS-22);江苏科技大学研究生科技创新计划项目(No.KYLX_1077);江苏省自然科学基金青年基金项目(No.BK2012273)

摘　　要：在之前的试验中给家蚕多化性品种马富(MF)的蛹体注射滞育激素仍无法诱导其产下滞育卵,为探究其原因,从该品种5龄第3天幼虫头部组织克隆了滞育激素基因(Bm DH)的全长c DNA,BLAST比对显示该基因编码氨基酸序列与NCBI数据库中来自家蚕二化性品种大造的Bm DH氨基酸序列(Gen Bank登录号:BAA059713)的相似度为90.1%,但存在差异的序列不包含滞育激素作用的功能区域。运用半定量RT-PCR与实时荧光定量PCR技术检测Bm DH基因在3个不同化性家蚕品种蛹期头部和卵巢的转录水平存在差异,转录水平从高到低依次为二化性家蚕品种C108,多化性品种MF、尼斯塔里;Bm DH在多化性家蚕品种MF的5龄期雌性和雄性幼虫的头部、血液、生殖腺3个组织中有转录,在中肠、丝腺、马氏管、表皮、脂肪体5个组织中没有检测到转录。研究结果初步提示:家蚕无滞育现象可能与体内滞育激素不足有关;但多化性品种MF不能被蛹期体外注射滞育激素诱导产生滞育卵与Bm DH基因的转录表达水平并无直接的关系。In order to explore why silkworm （Bombyx mori） multivoltine variety MF could not lay diapause eggs after in- jection of diapause hormone （DH） into pupa as shown in our previous experiment, the full length cDNA sequence of Bm- DH gene was cloned from head tissue of day 3 larvae of the fifth instar of MF. BLAST alignment showed that the amino sequence encoded by BmDH has 90. 1% sequence identity with the amino acid sequence of BmDH from silkworm bivolt- ine variety Dazhao （GenBank accession No. BAA059713）, but the different sequence did not contain the functiona） domain of diapause hormone. Semi-quantitative RT-PCR and real time fluorescent quantitative PCR indicated that the transcriptional level of BmDH gene in head and ovary of 3 silkworm varieties with different voltinisms at pupal stage was different, and was in the order of C108 （bivoltine variety） 〉MF （ multivoltine variety） 〉Nistari （multivoltine variety）. BmDH gene was expressed in head, blood and gonad of the 5th instar female and male larvae of multivoltine variety MF, while it could not be detected in midgut, silkgland, Malpighian tubule, epidermis and fat body. These results suggested that there is a certain relationship between non-diapause and diapause hormone deficiency in silk- worm. However, silkworm multivoltine variety MF cannot lay diapause eggs after injection of diapause hormone,indicating that it has no direct relationship with the transcription level of DH gene.

关 键 词：家蚕滞育激素基因 多化性品种 组织表达 实时荧光定量PCR 

分 类 号：Q786[生物学—分子生物学] S881.24[农业科学—特种经济动物饲养]