中华蜜蜂apiscerana cerana lsd-1基因的克隆及其表达特性分析-cloning and expression analysis of apitheraria ce rana lsd - 1 gene from apitheraria cerana.docxVIP

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中华蜜蜂apiscerana cerana lsd-1基因的克隆及其表达特性分析-cloning and expression analysis of apitheraria ce rana lsd - 1 gene from apitheraria cerana.docx

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中华蜜蜂apiscerana cerana lsd-1基因的克隆及其表达特性分析-cloning and expression analysis of apitheraria ce rana lsd - 1 gene from apitheraria cerana

中华蜜蜂(Apis cerana cerana) LSD-1 基因的克隆及其表达特性分析 PCR. Using the analysis soft TFBIND, we find that in addition to the typical TATA-box and CAAT-box, the binding sites of PPARγ and CAAT/enhancer binding protein α (C/EBPα), which are the major and determining adipogenic transcription factors, were found in this region. In addition, several important transcription factors such as heat shock factors (HSF) and T-cell factor required for regulating various environmental stresses were predicted. 2. The expression profile of AccLSD-1 at the different developmental stages was detected by real-time quantitative PCR. The results indicated that AccLSD-1 constitutively expressed throughout feeding larval, pupal and adult stages. In the larval feeding stages (L3-L5), the AccLSD-1 transcripts showed a very steep growth as increasing ages of larvae. In adult stage, the expression of AccLSD-1 exhibited much stronger in youth workers (25 day-old) than childhood (2 day-old) and old age ones (50 day-old). Howerver, AccLSD-1 was transcribed in a parabola fashion rather than persistently reduced trend in nonfeeding pupal stage and exhibt a highest level in brown-eyed pupae (Pb) stage. Moreover, there was an abrupt decrease in the quantity of transcripts in both larval-pupal transition and pupal-adult transition. The effects of various concentrations of conjugated linoleic acid (CLA) or rosiglitazone (Rosi) diets on the expression pattern of AccLSD-1 were investigated through rearing honeybee in laboratory. The results revealed that the transcripts of AccLSD-1 could be up-regulated by Rosi and down-regulated by CLA. The congenerous application of CLA and Rosi showed obvious alteration of the AccLSD-1 expression that was significantly higher than that of CLA alone group. A 600 aa 5′fragment of AccLSD-1 was selected to construct an E.coli expression vector pET- AccLSD-1-600, and then it was induced by IPTG to express in E.coli strain BL21 (DE3). The strong induced fusion protein bands were coll