SYBR Green Ⅰ实时荧光PCR在染色体常见非整倍体诊断中的应用  

Clinical application of SYBR Green Ⅰ real-time fluorescence quantitative PCR for diagnosis of common chromosome aneuploidy

作　　者：刘丽娟[1] 熊丽[2] 刘洁[2] 邓康[2] 刘思平[2] 吴瑞枫[2] 贾蓓[2] 宋兰林[2] 钟梅[2] 曾嵘[1] 

机构地区：[1]南方医科大学基础医学实验教学中心,广东广州510515 [2]南方医科大学南方医院妇产科,广东广州510515

出　　处：《南方医科大学学报》2010年第1期11-15,共5页Journal of Southern Medical University

基　　金：广东省科技攻关项目(2009B030801228);南方医科大学南方医院新业务新技术课题(2007030)

摘　　要：目的探讨SYBRGreenⅠ实时荧光PCR技术用于非整倍体诊断的可行性。方法选择13号染色体上的ABCC4基因、18号染色体上的TYMS基因、21号染色体上的DSCR3基因、X染色体上的HPRT2基因和Y染色体上的SRY基因为目的基因,以12号染色体上的管家基因GAPDH为参照基因,选取SYBRGreenⅠ作为荧光染料,采用双标准曲线相对定量PCR法进行检测,并与核型分析结果进行比对。结果检测13号染色体时,计算目的基因与参照基因比值,正常标本组(0.90±0.31)与三体标本组(1.39±0.12)的差别有显著意义(P=0.003);检测18号染色体时,正常标本组(1.07±0.44)与三体标本组(1.66±0.12)的差别有显著意义(P=0.000);检测21号染色体时,正常标本组(0.84±0.27)与三体标本组(1.73±0.54)的差别有显著意义(P=0.000);检测X染色体时,45,X组(0.62±0.12)与46,XY组(0.63±0.25)比值无差别(P=0.965),46,XX组(1.32±0.37)与47,XXY组(1.20±0.35)比值无差别(P=0.326),单个拷贝X(包括45,X、46,XY)(0.63±0.23)与两个拷贝X(46,XX、47,XXY)(1.26±0.36)比值差别有统计学意义(P=0.000);检测Y染色体时,正常女性(46,XX)无目的基因扩增,正常男性组(46,XY)(1.57±0.54)与三体标本组(47,XYY)(3.08±0.15)比值差别有统计学意义(P=0.003)。结论SYBRGreenⅠ实时荧光定量技术可用于快速诊断染色体非整倍体。Objective To investigate the value of real-time fluorescence quantitative PCR in the diagnosis of chromosome anepuploidy. Methods ABCC4 gene on chromosome 13, TYMS gene on chromosome 18, DSCR3 gene on chromosome 21, HPRT2 gene on chromosome X, and SRY gene on Y chromosome were used as the target genes, with GAPDH gene on chromosome 12 as the control gene. Using double-standard curve fluorescent relative quantitative PCR method with SYBR Green as the fluorescent dye, the gene expression levels were detected and the results were compared with those of karyotype analysis. Results The ratio of the target gene on chromosome 13 to the control gene showed a significant difference between the normal karyotype group （0.90±0.31） and trisome group （1.39±0.12, P=0.003）, and the genes on chromosome 18 （1.07± 0.44 vs 1.66±0.12, P=0.000） and chromosome 21 （0.84±0.27 vs 1.73±0.54, P=0.000） showed similar results. The expression of the genes on the X chromosome showed no significant difference between 45, X group and 46,XY group （0.62±0.12 vs 0.63± 0.25, P=0.965）, nor between 46, XX group and 47,XXY group （1.32±0.37 vs 1.20±0.35, P=0.326）, while a significant difference was noted between the single copy X （including 45,X and 46,XY） and two copies X （46,XX and 47,XXY） （0.63± 0.23 vs 1.26±0.36, P=0.000）. The expression of the target gene on the Y chromosome was not detected in normal females （46, XX）, and a significant difference in the expression was found between normal male group （46,XY） and 47,XYY group （1.57± 0.54 vs 3.08±0.15, P=0.003）. Conclusion SYBR Green I real-time fluorescence quantitative PCR can be used for the purpose of rapid diagnosis of chromosome aneuploidy.

关 键 词：非整倍体 实时荧光PCR SYBR Green Ⅰ 

分 类 号：R714.5[医药卫生—妇产科学]