YFP-H148Q/I152L在FRT细胞中的表达及其对碘离子敏感特性的研究  

Expression of YFP-H148Q/I152L in FRT cells and investigation of iodine ion sensitivity

作　　者：郑锴[1] 朱杭飞 王长文[2] 郭素红[1] 李正祎[1] 藏雨轩[1] 张雲乔 郑岩[1] 方芳[1] 郝峰[1] 

机构地区：[1]吉林医药学院检验学院,吉林吉林132013 [2]吉林医药学院公共卫生学院,吉林吉林132013

出　　处：《中国兽医杂志》2015年第10期11-13,I0001,共4页Chinese Journal of Veterinary Medicine

基　　金：吉林省教育厅基金资助课题(2013-351);2014年吉林省大学生创新创业训练计划;2013年吉林省大学生创新创业训练计划;吉林医药学院大学生科研基金资助课题(吉医学科字[2012]第12号);国家自然科学基金资助项目(81202031)

摘　　要：为探讨YFP-H148Q/I152L在FRT细胞中的表达及其对碘离子的敏感性,应用点突变试剂盒将真核表达载体YFP特异编码序列第148位氨基酸H突变为Q,第152位氨基酸I突变为L,脂质体介导将YFP-H148Q/I152L转染入稳定表达Ano2的FRT细胞中,应用荧光淬灭动力学试验检测YFP-H148Q/I152对其碘离子的敏感性。测序结果证实,成功将YFP突变为YFP-H148Q/I152L,且荧光淬灭动力学试验表明,表达YFP-H148Q/I152L的FRT细胞在加入激活剂和碘离子后,相对荧光强度显著下降。表明成功构建YFP-H148Q/I152L真核表达载体,并证实表达于FRT胞浆中的YFPH148Q/I152L具有对碘离子敏感的特性。To investigate the expression of YFP-H148Q/I152 L in FRT cells and its property to iodine ion sensitivity.Methods：The QuickChange Site-Directed Mutagenesis Kit was applied and then the YFP-pcDNA3.1 of the 148 th amino acids as H was mutated into Q and the 152 th amino acids as 1 was mutated into L.The eukaryotic expression vectors of YFP-H148Q/1152 L were transfected into FRT cells with expressed Ano2 by liposome.The iodine ion sensitivity of YFP-H148Q/1152 L was detected by the test of kinetics of fluorescence quenching.Results：The results of sequencing indicated that YFP was mutated into YFP-H148Q/I152 L.The results of the test of kinetics of fluorescence quenching indicated that in FRT cells added activator and iodine ion,the relative fluorescence intensity of YFP-H148Q/I152 L was significantly decreased.Conclusions：The eukaryotic vector of YFPH148Q/I152 L was successfully established and it was confirmed that the expressed YFP-H148Q/I152 L in FRT cytoplasms was sensitive to iodine ion.

关 键 词：YFP-H148Q/I152L FRT细胞 相对荧光强度 

分 类 号：Q785.786[生物学—分子生物学] Q256