拟穴青蟹chh基因近端启动子的功能分析
发布时间：2018-12-11 06:01
【摘要】：甲壳动物高血糖激素家族在调控甲壳动物的变态发育、生殖、蜕皮、血糖平衡及体内的渗透压平衡等方面起着重要的作用,甲壳动物高糖激素(CHH)是甲壳动物高血糖激素家族中的重要成员.通过Tail-PCR方法克隆获得拟穴青蟹(Scylla paramamosain)chh基因的近端启动子区序列,从翻译起始位点(ATG)起,共657 bp.生物信息学分析结果显示,转录起始位点位于翻译起始位点(ATG)上游第74个碱基(A),在转录起始位点(A)上游31 bp处存在TATA box;核心启动子区位于-41~+9 bp之间,包含在线软件预测的转录起始位点;潜在的转录因子结合位点包括Oct-1、C/EBPalp、GATA-1、Sp1、Ap-1、NF-κappa B、NF-1、RAP1、HNF-3等;目前获得的chh基因启动子区中没有Cp G岛.成功构建了chh基因4个启动子不同长度片段C1—C4,分别位于-583~+205、-388~+205、-286~+205、-46~+205 bp.含启动子不同长度片段的表达载体瞬时转染HEK293T细胞和Sf9细胞后的活性分析表明,在-286~-46、-583~-388 bp区域可能存在负调控基因表达的转录因子结合位点,在-388~-286 bp区域可能存在正调控基因表达的转录因子结合位点.
[Abstract]:The crustacean hyperglycemic hormone family plays an important role in regulating crustacean metamorphosis, reproduction, molting, blood glucose balance and osmotic balance in vivo. Crustacean hyperglycemic hormone (CHH) is an important member of the crustacean hyperglycemic hormone family. The proximal promoter sequence of (Scylla paramamosain) chh gene of Scylla sinensis was cloned by Tail-PCR method. The promoter sequence was 657 bp. from the translation initiation site (ATG). Bioinformatics analysis showed that the transcriptional initiation site was located at the 74th base (A), upstream of the translation initiation site (ATG) and TATA box; existed at 31 bp upstream of the transcription initiation site (A). The core promoter is located between -41 ~ 9 bp and contains the transcription initiation site predicted by online software. The potential transcription factor binding sites include Oct-1,C/EBPalp,GATA-1,Sp1,Ap-1,NF- 魏 appa BF-1, RAP1, HNF-3 and so on. There is no Cp G island in the promoter region of chh gene. Four promoters of chh gene, C1-C4, were successfully constructed and located at -583-205 bp.. The activity analysis of the expression vector containing different length of promoter in HEK293T cells and Sf9 cells showed that there might be transcription factor binding sites in the bp region of -286G -46-46-586 -583 -388 bp. There may be transcription factor binding sites in the region of-388- 286 bp that positively regulate gene expression.
【作者单位】： 集美大学水产学院农业部东海海水健康养殖重点实验室;福建农林大学动物科学学院;
【基金】：国家自然科学基金资助项目(41176152) 集美大学创新团队基金资助项目(2010A001)
【分类号】：S917.4;Q78
本文编号：2372012
[Abstract]:The crustacean hyperglycemic hormone family plays an important role in regulating crustacean metamorphosis, reproduction, molting, blood glucose balance and osmotic balance in vivo. Crustacean hyperglycemic hormone (CHH) is an important member of the crustacean hyperglycemic hormone family. The proximal promoter sequence of (Scylla paramamosain) chh gene of Scylla sinensis was cloned by Tail-PCR method. The promoter sequence was 657 bp. from the translation initiation site (ATG). Bioinformatics analysis showed that the transcriptional initiation site was located at the 74th base (A), upstream of the translation initiation site (ATG) and TATA box; existed at 31 bp upstream of the transcription initiation site (A). The core promoter is located between -41 ~ 9 bp and contains the transcription initiation site predicted by online software. The potential transcription factor binding sites include Oct-1,C/EBPalp,GATA-1,Sp1,Ap-1,NF- 魏 appa BF-1, RAP1, HNF-3 and so on. There is no Cp G island in the promoter region of chh gene. Four promoters of chh gene, C1-C4, were successfully constructed and located at -583-205 bp.. The activity analysis of the expression vector containing different length of promoter in HEK293T cells and Sf9 cells showed that there might be transcription factor binding sites in the bp region of -286G -46-46-586 -583 -388 bp. There may be transcription factor binding sites in the region of-388- 286 bp that positively regulate gene expression.
【作者单位】： 集美大学水产学院农业部东海海水健康养殖重点实验室;福建农林大学动物科学学院;
【基金】：国家自然科学基金资助项目(41176152) 集美大学创新团队基金资助项目(2010A001)
【分类号】：S917.4;Q78
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