牛樟芝鲨烯合酶基因的克隆与表达分析The Cloning and Expression Analysis of Squalene Synthetase Gene(AcSQS) in Antrodia camphorata
原晓龙;赵能;华梅;陈剑;杨宇明;王娟;王毅;
摘要(Abstract):
为了解牛樟芝三萜类生物合成、调控机理,以多孔菌科真菌茯苓(Wolfiporia cocos,AFR13032.1)的SQS基因为模板对牛樟芝基因组数据进行本地Blast,分离获得牛樟芝鲨烯合酶(AcSQS)基因.以该序列为模板设计特异引物Ac SQSF0和Ac SQSR0,通过PCR扩增得到Ac SQS c DNA全长,并对其进行生物信息学分析,检测不同碳氮源添加物的培养基上该基因的表达水平.结果显示:克隆得到的基因为牛樟芝鲨烯合酶(Ac SQS),该基因含4个外显子,3个内含子,外显子拼接总长1 803 bp,编码600个氨基酸;其蛋白序列的117位为信号肽位点,具两段跨膜结构,可能定位于内质网;位于该蛋白18917位为信号肽位点,具两段跨膜结构,可能定位于内质网;位于该蛋白189204位的CHYVAGLVGEGLTRL和225204位的CHYVAGLVGEGLTRL和225238位的MGLMLQKTNIIRDY的保守基序为鲨烯合酶识别位点,DTIEDD和D(Y/F)RED为FPP绑定位点;蛋白网络分析显示其位于三萜类化合物的代谢网络中;聚类分析显示Ac SQS蛋白和茯苓、硫磺多孔菌、拟蜡菌、灵芝、云芝等的SQS蛋白聚为一支;不同碳氮源添加物培养基上的表达分析显示:果糖诱导该基因表达的能力最强,而不同氮源添加物诱导该基因表达的能力无明显差异.
关键词(KeyWords): 牛樟芝;鲨烯合酶;基因克隆;表达分析
基金项目(Foundation): 对外科技合作计划项目(2015IA004);; 国家自然科学基金青年项目(31400488);; 云南省应用基础研究计划面上项目(2016FB055)
作者(Authors): 原晓龙;赵能;华梅;陈剑;杨宇明;王娟;王毅;
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