乳腺癌klk6基因表达的实时荧光定量法的建立及初步应用  

Development and primary application of a real-time quantitative polymerase chain reaction for the detection of human breast cancer related novel gene-kallikrein gene 6 expression

作　　者：胡成进[1] 沈茜[1] 陈英剑[2] 闻新棉[3] 

机构地区：[1]上海第二军医大学长海医院实验诊断科,200433 [2]济南军区总医院实验诊断科 [3]山东大学生命科学院

出　　处：《中华检验医学杂志》2005年第6期610-612,共3页Chinese Journal of Laboratory Medicine

摘　　要：目的建立人乳腺癌相关基因组织型激肽释放酶基因家族成员Kallikrein6(klk6)mRNA荧光定量逆转录聚合酶链反应(FQPCR)检测方法,以检测正常乳腺和乳腺癌组织中klk6基因表达水平。方法以GAPDH作参照,用SYBRGreenI荧光定量技术,建立检测klk6mRNA的FQPCR方法;并检测32份正常乳腺和乳腺癌组织标本的循环阈值(Ct),然后,通过REST软件分析klk6mRNA的表达水平。结果FQPCR方法对GAPDH和klk6mRNA的扩增效率分别为0.90和0.95,批内变异系数(CV)分别为1.0%～2.1%和0.8%～1.2%,批间CV分别为3.2%和3.9%。正常乳腺和乳腺癌组织的klk6对GAPDH的相对表达量分别为0.017±0.009和0.040±0.017。用REST软件分析,提示乳腺癌组织klk6表达水平上调。结论建立的klk6mRNAFQPCR检测方法简便,特异,重复性好,可信度高,可用于klk6基因表达水平与肿瘤相关性研究。Objective To develop a real-time quantitative PCR method for detection of human breast cancer related novel gene-kallikrein gene 6 (klk6) expression and investigate klk6 expression levels in breast cancer tissue.Methods Using Sybr Green I, with GAPDH as reference, a real-time quantitative PCR method was established. klk6 expression levels of 32 normal and breast cancer tissues were detected and analyzed by the method and REST software.Results The amplification efficiencies for GAPDH and klk6 of the real-time quantitative PCR method were 0.90 And 0.95, respectively; inter-coefficient of variation were 1.0%~2.1% and 0.8%~1.2%,respectively; intra-coefficient of variation were 3.2% and 3.9%, respectively. The relative expression levels of klk6 in normal breast and breast cancer tissues were 0.017±0.009 and 0.040±0.017 with GAPDH as reference. Analysis results with REST indicated klk6 expression was up- regulated in breast cancer.Conclusion The real-time quantitative PCR method with Sybr Green I for klk6 mRNA quantification was simple, specific, reproductive and reliable, and could be used to study relationship betweeen klk6 expression and tumor.

关 键 词：实时荧光定量法 荧光定量逆转录聚合酶链反应 初步应用 变异系数(CV) 基因表达水平 乳腺癌组织 GAPDH 乳腺癌相关基因 Q-PCR方法 PCR检测方法 mRNA 正常乳腺 荧光定量技术 软件分析 Green 癌组织标本 相关性研究 家族成员 

分 类 号：R737.9[医药卫生—肿瘤]