【摘要】：甲殼動物高血糖激素家族在調(diào)控甲殼動物的變態(tài)發(fā)育、生殖、蛻皮、血糖平衡及體內(nèi)的滲透壓平衡等方面起著重要的作用,甲殼動物高糖激素(CHH)是甲殼動物高血糖激素家族中的重要成員.通過Tail-PCR方法克隆獲得擬穴青蟹(Scylla paramamosain)chh基因的近端啟動子區(qū)序列,從翻譯起始位點(ATG)起,共657 bp.生物信息學(xué)分析結(jié)果顯示,轉(zhuǎn)錄起始位點位于翻譯起始位點(ATG)上游第74個堿基(A),在轉(zhuǎn)錄起始位點(A)上游31 bp處存在TATA box;核心啟動子區(qū)位于-41~+9 bp之間,包含在線軟件預(yù)測的轉(zhuǎn)錄起始位點;潛在的轉(zhuǎn)錄因子結(jié)合位點包括Oct-1、C/EBPalp、GATA-1、Sp1、Ap-1、NF-κappa B、NF-1、RAP1、HNF-3等;目前獲得的chh基因啟動子區(qū)中沒有Cp G島.成功構(gòu)建了chh基因4個啟動子不同長度片段C1—C4,分別位于-583~+205、-388~+205、-286~+205、-46~+205 bp.含啟動子不同長度片段的表達載體瞬時轉(zhuǎn)染HEK293T細胞和Sf9細胞后的活性分析表明,在-286~-46、-583~-388 bp區(qū)域可能存在負調(diào)控基因表達的轉(zhuǎn)錄因子結(jié)合位點,在-388~-286 bp區(qū)域可能存在正調(diào)控基因表達的轉(zhuǎn)錄因子結(jié)合位點.
[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.
【作者單位】： 集美大學(xué)水產(chǎn)學(xué)院農(nóng)業(yè)部東海海水健康養(yǎng)殖重點實驗室;福建農(nóng)林大學(xué)動物科學(xué)學(xué)院;
【基金】：國家自然科學(xué)基金資助項目(41176152) 集美大學(xué)創(chuàng)新團隊基金資助項目(2010A001)
【分類號】：S917.4;Q78

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本文編號：2372012