葱蝇Dorsal基因的克隆·表达及生物信息学分析

目的]探讨Dorsal与葱蝇滞育发育的关系。方法]以葱蝇(Delia antiqua)为试虫,采用RACE方法克隆了葱蝇Dorsal的全长cDNA序列;将该序列推导的氨基酸序列在GenBank搜寻同源序列,与搜寻到的14个代表性昆虫Dorsal序列进行相似性比较,并进行系统进化分析;采用半定量RT-PCR方法分析了Dorsal基因在葱蝇非滞育、夏滞育、冬滞育蛹中的表达情况。结果]克隆了葱蝇Dorsal基因的cDNA序列,全长2 412 bp,开放阅读框1 974 bp,编码657个氨基酸,等电点PI约8.5,分子量72.9 kDa。与GenBank中的其他14个昆虫代表性Dorsal序列相似性比较和进化分析显示葱蝇的Dorsal与黑腹果蝇(Drosophila melanogaster)、拟暗果蝇(Drosophlia pseudoob-scura)的Dorsal序列相似性最大。半定量分析显示葱蝇Dorsal基因在葱蝇冬滞育、夏滞育、非滞育蛹滞育发育关键期的表达量均明显升高,尤其在滞育后期表达量最高,初步判断Dorsal基因与葱蝇滞育发育相关。结论]为进一步开展Dorsal基因在昆虫滞育发育过程中的功能研究奠定了基础。

Cloning,Expression and Bioinformatic Analysis of Dorsal Gene in Delia antiqua

Objective] The aim was to explore the relationship between the Dorsal and diapause development of Delia antiqua.Method] The full-length cDNA of Dorsal in D.antiqua was cloned through RACE.The similarity comparison among deduced amino acid sequence of DaDorsal cDNA and the Dorsals of other 14 insect species was compared,and the phylogenetic analysis of these Dorsals was conducted.The expression level of Dorsal gene in winter-,summer-and non-diapausing pupae was analyzed.Result] The full-length of Dorsal cDNA sequence,2 412 bp,was obtained with ORF 1 974 bp,which coded 657 amino acids with predicted Mw 72.9 kDa and PI 8.5.The result of similarity comparison indicated that the DaDorsal was most related to those of Drosophlia melanogaster and Drosophlia pseudoobscura.The result of semi-quantitative RT-PCR analysis showed the expression level of Dorsal gene increased in characteristic duration of winter-,summer-and non-diapausing pupae,eapecially at the late diapause,which might imply its relationship to D.antiqua diapause and development.Conclusion] The study lays the foundation for further study on gene function of Dorsal in insect diapause and development.