双链探针同步荧光技术快速筛查C282Y点突变.pdfVIP

遗 　传 H ER EDI TA S ( B eij ing) 25 ( 1) :9～13 ,2003 　　　　　　　　　　　　　　　　　　　　　　　　研究报告 双链探针同步荧光技术快速筛查 C2 82 Y 点突变 张永有 ,李庆阁 ,栾国彦 ,梁基选 ( 厦门大学生命科学学院细胞生物学与肿瘤细胞工程教育部重点实验室 ,厦门 36 1005) 摘 　要 : 以荧光染料 Fam 和Joe 分别标记野生型和突变型双链探针作为均相检测探针 , 以构建的 DNA 模板作为研 究模型 ,采用固定波长差同步荧光分析法对 PCR 反应产物进行终点检测 。通过对 H FE 基因 C282 Y 点突变的检 测 ,并以限制性内切核酸酶 Rs a I 证实 ,该方法是一种廉价 、快速 、可靠的筛查遗传性血色病基因 C282 Y 突变的方 法 ,该法可扩展到各种基因的突变检测 。 关键词 :遗传性血色病 ;双链探针 ; 同步荧光 ;基因突变筛查 中图分类号 :Q3 - 3 　　　文献标识码 :A 　　　 文章编号 :0253 - 9772 (2003) 0 1 - 0009 - 05 Screening C282 Y Mutation with DoubleStranded Probes Using Synchronous Fluorometry ZHAN G YongYou ,L I QingGe ,L UAN GuoYan ,L IAN G J iXuan ( The Key L aboratory of Cell B iology an d T u mor Cell Engi neeri ng of The M i nist ry of Ed ucation , S chool of L if e S ciences , X iamen University , X iamen , Fuj ian 36 1005 , Chi na) Abstract :We described in t he p ap er a new hight hroughput screening met hod for Cys282 Tyr mutation in hereditary haemochromatosis wit h doublestranded probe using synchronous fluorometry . The probe for wild typ e was labeled wit h Fam ,t he probe for mutant typ e was labeled wit h Joe . After PCR ,reaction tubes were transferred to a sp ectrofluorometer , where synchronous sp ectra were scanned in a constantwavelengt h mode . The genotyp e could be obtained t hrough t he ap p earance of t he fluorescence p eaks corresponding to each probe . The result s were totally in agreement wit h restriction en donuclease analysis. Considering t he simplicity ,low cost and sp ecificity ,t his approach could be generally applied to detect varieties of gene mutations.