巴氏杜氏藻八氢番茄红素合成酶侧翼调控序列的克隆与分析

研究表明八氢番茄红素合成酶( PSY)是巴氏杜氏藻(Dunaliella bardawil)类胡萝卜素代谢途径中的第一个关键调节酶.本实验采用基于LAoPCR的基因组步移法分别设计两组基因特异引物pSP1-3及tSP1-3,克隆巴氏杜氏藻PSY( DbPSY)的启动子和终止子序列,并利用在线启动子分析软件分析其保守调控序列.最后利用生物信息学方法分析预测DbPSY的蛋白质结构.分析结果表明,DbPSY启动子具有两个保守调控序列:BOXLCOREDCPAL和GT1GMSCAM4,其中BOXLCOREDCPAL受紫外光诱导,上调下游基因的表达,GT1GMSCAM4受盐调控,促进下游基因的表达.蛋白结构分析表明,DbPSY的N端在邻近物种间具有较大的多样性.我们推测DbPSY的微调与启动子保守序列及蛋白质N端序列的多样性相关.

In silico Analysis of Phytoene Synthase and its Promoter Reveals Hints for Regulation Mechanisms of Carotenogenesis in Duanliella bardawil

Previous researches showed that phytoene synthase(PSY) is the first regulatory point in carotenogenesis in Dunaliella bardawil.In this study,two sets of adjacent gene specific primers(pSP1,pSP2 and pSP3;tSP1,tSP2 and tSP3) were designed and LA PCR-based genomic walking approach was performed for isolation of psy promoter and terminator in D.bardawil,respectively;and subsequent analysis of conserved regulatory sequences were conducted by online promoter analysis softwares.Finally,using bioinformatics methods,the protein structure of DbPSY was also predicted.The calculated results found two conserved regulatory sequences in PSY promoter: BOXLCOREDCPAL and GT1GMSCAM4.BOXLCOREDCPAL was induced by UV to up-regulate expression of downstream gene;and GT1GMSCAM4 up-regulates expression of downstream gene induced by NaCl.Protein structure analysis demonstrated low homology of N-terminus between D.bardawil and other algae.Therefore we speculated the regulation mechanisms may partly involve in the conserved sequences of psy promoter and the diversity of N-terminus of DbPSY compared to other species.