中文摘要：

【目的】昆虫变态发育的启动主要受前胸腺合成分泌的蜕皮激素所调控，而蜕皮激素的合成是由细胞色素P450基因催化完成。家蚕（Bombyx mori）是一种产丝昆虫，蚕业生产中如果能阻断或延迟家蚕蛹变态发育进程，将有利于改进蚕茧处理工艺，提高蚕丝品质。论文旨在鉴定参与家蚕蜕皮激素合成的细胞色素P450基因，从而为人为遗传调节家蚕变态发育提供靶基因。【方法】基于序列同源性比对，筛选家蚕及其他昆虫中参与蜕皮激素合成的P450基因。利用ClustalW软件，分析昆虫蜕皮激素合成相关P450基因的遗传发生关系。通过全基因组表达芯片数据分析及RT-PCR验证，调查家蚕蜕皮激素合成相关P450基因的时空表达特征。利用RNAi技术分析Cyp314a1表达下调对家蚕变态发育的影响。【结果】家蚕基因组中有4个参与家蚕蜕皮激素合成的P450基因，即Cyp306a1、Cyp302a1、Cyp315a1和Cyp314a1。比较分析显示，这4个蜕皮激素合成相关的P450基因在家蚕及其他昆虫中都是单拷贝，而且每个基因的同源体在遗传发生树上能很好地聚成一类，表明昆虫蜕皮激素合成相关的P450基因及其负责的蜕皮激素合成通路非常保守。时空表达谱分析显示，在家蚕幼虫5龄第3天，Cyp302a1、Cyp315a1和Cyp314a1主要在家蚕幼虫卵巢、精巢和头部等组织器官中高表达；在幼虫-蛹-成虫变态发育进程中，Cyp302a1、Cyp315a1和Cyp314a1主要在蛹变态发育后期表达，其中Cyp314a1分别在上簇、化蛹及羽化前高表达，这与蜕皮激素滴度高峰出现的时期基本一致。Cyp314a1的RNAi导致家蚕不能正常化蛹及雌蛾卵巢发育异常，且降低了蜕皮激素信号通路关键基因HR3及Ftz-f1的表达。【结论】家蚕蜕皮激素合成相关的P450基因在进化上非常保守，其表达水平降低能引起家蚕蛹变态发育受阻，暗示蜕皮激素合成相关P450基因可以用?

英文摘要：

【Objective】Insect metamorphosis is initiated by ecdysone, which is synthesized in prothoracic gland through cytochrome P450 enzymes-catalyzed enzymatic reactions. Silkworm （Bombyx mori） is a silk-producing insect. Interrupting or delaying the pupa-adult transition in silkworm will help to reform the flow of silkworm cocoon-processing and to further improve the quality of silk. The objective of this study is to identify and characterize P450 genes involving in ecdysteroidogenesis in silkworm and to provide candidate genes for genetic controlling of silkworm metamorphosis. 【Method】 Ecdysteroidogenesis-related P450 genes were identified in silkworm and other insects through sequence homology search. Phylogenetic tree of insect ecdysteroidogenesis- related P450 genes was constructed by ClustalW program. The mRNA expression profiles of ecdysteroidogenesis-related P450 genes from silkworm were checked by microarray and RT-PCR analysis. RNAi experiment was used to examine the effects of down- regulation of expression level of ecdysteroidogenesis-related P450 genes on silkworm metamorphosis. 【Result】Four P450 genes involving in ecdysteroidogenesis, including Cyp306a1, Cyp302a1, Cyp315a1, and Cyp314a1, were identified in the silkworm genome. Comparative analysis revealed that each edysteroidogenesis-related P450 gene existed as a single copy in the surveyed insects and all its orthologs in insects were grouped together well in the phylogenetic tree, indicating edysteroidogenesis-related P450 genes and edysteroidogenesis pathway were very conserved during insect evolution. Spatio-temporal expression profiling found that Cyp302a1, Cyp315a1, and Cyp314a1 showed a high expression in the ovary, testis, and head of silkworm larvae and were continually expressed during the late stages of silkworm metamorphosis. Cyp314a1 expression could be highly detected before wandering, pupation, or eclosion, showing a consistency with the stage-specific changes of ecdysone pulses. RNAi of Cyp314a1 resulted in not only larval