家蚕无鳞片翅突变体scaleless产生机理的研究-特种经济动物饲养专业论文.docx

个原神经基因中BIn-ASHl是最早分化形成的一个；Bm-A跚2利Bm-ASH3同源性最高，推测相对 个原神经基因中BIn-ASHl是最早分化形成的一个；Bm-A跚2利Bm-ASH3同源性最高，推测相对 于Bm-ASHl形成较晚。用RT-PCR和实时荧光定量PCR检测了家蚕ASH基因在幼虫期不同组织 器官及胚胎发育过程中的表达情况，发现Bm-ASHl和BIn-ASH2表达范围最』“，胚胎发育过程中 也有较高的表达量；而Bm-ASH3表达特异性相对较高，因此推断其形成较Bm-ASH2为晚，即 Bm_ASH3是四个家蚕ASH基因中最晚形成的一个。ase基因主要在神经前体细胞中表达，冈此 Bm．ase表达范围较窄，组织中总体表达量较低。但同源比较和功能分析都不能将家蚕的三个原神 经基因和果蝇的一一对应，囚此其命名依然按照发现的先后顺序进行。 通过半定量RT-PCR发现家蚕的吐丝期利蛹期，BmMSHl和Bm以SH3基因在scaleless和 、帆1家蚕中的表达时相几乎一致：Bm-ASH2只在0 d～2 d的wT蛹翅中表达，而scaleless中没有 检测到表达信号。原位杂交结果也证明BIn-ASH2在scaleless蛹翅中不能正常表达。将wT及 scaleless的Bm-ASH2基因起始密码子上游的启动子序列克隆并测序，发现scaleless的ATG上游 1 027 bp处有一段连续26bp的缺失突变。利用家蚕细胞瞬时表达系统分析了Bm-ASH2基闶启动 子活性，发现scaleless比wT低得多，只与截去了其缺失的26 bp的979 bp wT启动子片段的活 性相当。EMSA结果显示scaleless缺失的26 bp序列可以特异性结合某种核蛋白。遗传学实验则 证明26bp序列缺失与无鳞片翅突变表型紧密连锁。这些都说明scaleless的Bm-ASH2基因因启动 子突变而使其不能正常转录和表达。利用瞬时表达系统使BIn-ASH2基冈在scaleless早期蛹翅中 表达，可以使翅面鳞片明显增多，进一步说明Bm-ASH2是家蚕翅面鳞片产生的芙键因子。从而 初步阐明了家蚕无鳞片翅突变体scaleless产生的分子机理。 关键词：家蚕，scaleless突变体，翅原基，鳞片，产生机理 Ⅱ AbstractThe Abstract The coating of scales on the surface of the wings is a basic characteristic of lepidopteran adults．Wing scales have important physical function to Lepidoptera，and pattems formed by multicolored scales not only have specifically developmental mechanisms，but also are a key innovation during lepidopteran evolution．So lepidopteran wing scales are valuable for the studies of insect physiology,developmental biology,biologic evolution，and even pest contr01．In this thesis，we studied the silkworm scaleless wings mutant(scaleless，s1)on morphological，cell biological and molecular biological levels，and tried to clarify the mechanism of the mutant and offer references for the studies the development of lepidopteran wing scales． We carried out studies beginning with the development of the wing discs／wings，in size and shape， during the 5“instar larval and pupal period．We found that the wing disc grew slowly in the prophase of the larva，while very fast just before metamorphism．We also found that pupa was a period of the