实时荧光PCR探针法和DNA测序法应用于CYP2C9和VKORC1基因检测的对比研究
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摘要
目的:比较实时荧光PCR探针法和DNA测序法检测细胞色素氧化酶P450(CYP2C9)和维生素K环氧化物还原酶复合体1(VKORC1)基因多态性的一致性,建立适用于临床推广的华法林基因多态性检测的方法。方法:本研究随机收集清远市人民医院20例就诊患者的全血标本,应用实时荧光PCR探针法和DNA测序法检测其CYP2C9和VKORC1基因的多态性,结果运用Excel软件统计并进行比较分析。结果:两种方法对CYP2C9和VKORC1基因多态性的检测结果一致,且实时荧光PCR探针法比DNA测序法更简易、经济、快捷。结论:与DNA测序法相比,应用实时荧光PCR探针法检测CYP2C9和VKORC1基因的突变具有较高的临床应用价值,比较适用于临床检测。
Objective:To compare the polymorphism of cytochrome oxidase P450(CYP2C9) and vitamin K epoxide reductase complex 1(VKORC1) gene by real-time fluorescent PCR and DNA sequencing.Methods of clinical promotion of warfarin gene polymorphism.Method:The whole blood samples of 20 patients in Qingyuan People's Hospital were randomly collected.The polymorphisms of CYP2C9 and VKORC1 genes were detected by real-time fluorescent PCR and DNA sequencing.The results were analyzed by Excel software and compared. Result:The results of two methods were consistent with the detection of CYP2C9 and VKORC1 gene polymorphism,and the real-time fluorescent PCR method was simpler,more economical and faster than DNA sequencing method.Conclusion:Compared with DNA sequencing method,real-time fluorescent PCR probe method detection of CYP2C9 and VKORC1 gene mutations has a high clinical value,is more suitable for clinical testing.
Objective:To compare the polymorphism of cytochrome oxidase P450(CYP2C9) and vitamin K epoxide reductase complex 1(VKORC1) gene by real-time fluorescent PCR and DNA sequencing.Methods of clinical promotion of warfarin gene polymorphism.Method:The whole blood samples of 20 patients in Qingyuan People's Hospital were randomly collected.The polymorphisms of CYP2C9 and VKORC1 genes were detected by real-time fluorescent PCR and DNA sequencing.The results were analyzed by Excel software and compared. Result:The results of two methods were consistent with the detection of CYP2C9 and VKORC1 gene polymorphism,and the real-time fluorescent PCR method was simpler,more economical and faster than DNA sequencing method.Conclusion:Compared with DNA sequencing method,real-time fluorescent PCR probe method detection of CYP2C9 and VKORC1 gene mutations has a high clinical value,is more suitable for clinical testing.
引文
[1]Johnson J A.Warfarin pharmacogenetics:a rising tide for its clinical value[J].Circulation,2012,125(16):1964-1966.
[2]张引兰,张郁峰.基因多态性对华法林个体化药量作用的临床分析[J].实用药物与临床,2012,15(4):196-198.
[3]吴海丽,向道康,黄陵,等.SYBR GreenⅠ实时荧光PCR检测VKORC1和CYP2C9基因型[J].江苏医药,2011,37(15):1815-1818.
[4]吴薇,李艳,童永清,等.DNA测序法检测华法林用药剂量相关基因多态性的应用价值[J].中华检验医学杂志,2012,35(11):1016-1020.
[5]崔广林,丁虎,徐宇军,等.高分辨率熔解曲线方法在指导华法林使用起始剂量基因分型中的应用及临床各种基因分型方法的比较[J].中华心血管病杂志,2012,40(6):477-481.
[6]张亚同,梁欣,胡欣,等.基于连接酶反应快速检测华法林相关基因VKORC1、CYP2C9多态性[J].临床药理学,2010,15(12):1395-1401.
[7]Redman A R.Implications of cytochrome P450 2C9 polymorphisms on warfarin metabolism and dosing[J].Pharmacotherapy,2001,21(2):235-242.
[8]Lee C R,Coldstein J A,Pieper J A.Cytochrome P450 2C9polymorphisms a comprehensive review of the in-vitro and human data[J].Phammcogeneties,2002,12(3):251-263.
[9]Sullivan-Klose T H,Ghanayem B I,Bell D A,et al.The role of the CYP2C9-Leu359 allelic wariant in the tolbutamide polymorphism[J].Pharmacogenetics,1996,6(4):341-349.
[10]Sanderson S,Emery J,Higgins J.CYP2C9 gene variants,drug dose,and bleeding risk in warfarin-treated patients a Hu GEnet systematic review and meta-analysis[J].Genet Med,2005,7(2):97-104.
[11]Reitsma P H,van der Heijden J F,Groot A P,et al.AC1173T dimorphism in the VKORC1 gene determines coumarin sensitivity and bleeding risk[J].PLo S Med,2005,2(10):e312.
[12]柴树红,陈丽,谷苞,等.荧光PCR-熔解曲线法检测新疆回族人群CYP2C9和VKORC1基因多态性研究[J].国际检验医学杂志,2016,37(17):2371-2373.
[13]戎国栋,沈丹婷,陈丹,等.基于荧光定量PCR技术的人类CYP2C9和VKORC1基因多态性检测方法学评价[J].临床与病理杂志,2017,37(11):2357-2362.
[14]魏君,靳衡,柴艳芬,等.CYP2C9和VKORC1基因检测指导急性肺血栓栓塞症患者个体化华法林抗凝治疗的临床研究[J/OL].中华危重症医学杂志(电子版),2016,9(2):91-95.
[15]史长城,田港,吴静,等.CYP2C9和VKORC1基因检测指导华法林给药的有效性和安全性的Meta分析(英文)[J].中国新药与临床杂志,2015,34(11):850-857.
[16]吴薇,韩瑞玲,王丽岳,等.DNA测序法检测CYP2C19基因多态性指导氯吡格雷用药的临床价值[J].检验医学与临床,2015,12(4):433-435.
[17]栗冬梅,刘云彦,杜鹏程,等.实时荧光定量Taq Man-MGB探针法检测杆菌样巴尔通体[J].微生物学通报,2015,42(2):427-435.
[18]马莉,刁戈,肖小璞,等.DNA测序法检测血友病B患者F9基因突变[J].中国输血杂志,2013,26(11):1084-1087.
[19]高正琴,岳秉飞,关伟鸿.CAR菌Taq Man MGB探针法实时荧光定量PCR快速检测方法的建立与应用[J].中国比较医学杂志,2013,23(8):9-13,19.
[20]姜文灿,岳素文,江洪,等.分子信标-Taq Man探针法实时荧光定量PCR新型探针及引物[J].生物技术通报,2016,32(11):107-114.
[21]顾梦月,张希根,张楚麟,等.实时荧光定量PCR检测CYP2C9*3及VKORC1-C1173T基因多态性方法的建立[J].药物生物技术,2016,23(3):195-200.
[22]尹栩芳,蒋卫霞,饶品彬.LNA-taqman探针实时荧光PCR快速检测CYP4F2-C1347T,CYP2C9*3,VKORC1-C1173T与VKORC1-G1639A基因多态性方法的建立[J].现代检验医学杂志,2017,32(6):1-5.
[2]张引兰,张郁峰.基因多态性对华法林个体化药量作用的临床分析[J].实用药物与临床,2012,15(4):196-198.
[3]吴海丽,向道康,黄陵,等.SYBR GreenⅠ实时荧光PCR检测VKORC1和CYP2C9基因型[J].江苏医药,2011,37(15):1815-1818.
[4]吴薇,李艳,童永清,等.DNA测序法检测华法林用药剂量相关基因多态性的应用价值[J].中华检验医学杂志,2012,35(11):1016-1020.
[5]崔广林,丁虎,徐宇军,等.高分辨率熔解曲线方法在指导华法林使用起始剂量基因分型中的应用及临床各种基因分型方法的比较[J].中华心血管病杂志,2012,40(6):477-481.
[6]张亚同,梁欣,胡欣,等.基于连接酶反应快速检测华法林相关基因VKORC1、CYP2C9多态性[J].临床药理学,2010,15(12):1395-1401.
[7]Redman A R.Implications of cytochrome P450 2C9 polymorphisms on warfarin metabolism and dosing[J].Pharmacotherapy,2001,21(2):235-242.
[8]Lee C R,Coldstein J A,Pieper J A.Cytochrome P450 2C9polymorphisms a comprehensive review of the in-vitro and human data[J].Phammcogeneties,2002,12(3):251-263.
[9]Sullivan-Klose T H,Ghanayem B I,Bell D A,et al.The role of the CYP2C9-Leu359 allelic wariant in the tolbutamide polymorphism[J].Pharmacogenetics,1996,6(4):341-349.
[10]Sanderson S,Emery J,Higgins J.CYP2C9 gene variants,drug dose,and bleeding risk in warfarin-treated patients a Hu GEnet systematic review and meta-analysis[J].Genet Med,2005,7(2):97-104.
[11]Reitsma P H,van der Heijden J F,Groot A P,et al.AC1173T dimorphism in the VKORC1 gene determines coumarin sensitivity and bleeding risk[J].PLo S Med,2005,2(10):e312.
[12]柴树红,陈丽,谷苞,等.荧光PCR-熔解曲线法检测新疆回族人群CYP2C9和VKORC1基因多态性研究[J].国际检验医学杂志,2016,37(17):2371-2373.
[13]戎国栋,沈丹婷,陈丹,等.基于荧光定量PCR技术的人类CYP2C9和VKORC1基因多态性检测方法学评价[J].临床与病理杂志,2017,37(11):2357-2362.
[14]魏君,靳衡,柴艳芬,等.CYP2C9和VKORC1基因检测指导急性肺血栓栓塞症患者个体化华法林抗凝治疗的临床研究[J/OL].中华危重症医学杂志(电子版),2016,9(2):91-95.
[15]史长城,田港,吴静,等.CYP2C9和VKORC1基因检测指导华法林给药的有效性和安全性的Meta分析(英文)[J].中国新药与临床杂志,2015,34(11):850-857.
[16]吴薇,韩瑞玲,王丽岳,等.DNA测序法检测CYP2C19基因多态性指导氯吡格雷用药的临床价值[J].检验医学与临床,2015,12(4):433-435.
[17]栗冬梅,刘云彦,杜鹏程,等.实时荧光定量Taq Man-MGB探针法检测杆菌样巴尔通体[J].微生物学通报,2015,42(2):427-435.
[18]马莉,刁戈,肖小璞,等.DNA测序法检测血友病B患者F9基因突变[J].中国输血杂志,2013,26(11):1084-1087.
[19]高正琴,岳秉飞,关伟鸿.CAR菌Taq Man MGB探针法实时荧光定量PCR快速检测方法的建立与应用[J].中国比较医学杂志,2013,23(8):9-13,19.
[20]姜文灿,岳素文,江洪,等.分子信标-Taq Man探针法实时荧光定量PCR新型探针及引物[J].生物技术通报,2016,32(11):107-114.
[21]顾梦月,张希根,张楚麟,等.实时荧光定量PCR检测CYP2C9*3及VKORC1-C1173T基因多态性方法的建立[J].药物生物技术,2016,23(3):195-200.
[22]尹栩芳,蒋卫霞,饶品彬.LNA-taqman探针实时荧光PCR快速检测CYP4F2-C1347T,CYP2C9*3,VKORC1-C1173T与VKORC1-G1639A基因多态性方法的建立[J].现代检验医学杂志,2017,32(6):1-5.