非多态性位点多重荧光定量PCR技术在产前诊断胎儿染色体非整倍体异常中的应用  

Application of multiplex quantitative fluorescent PCR with non-polymorphic loci in prenatal diagnosis

作　　者：朱湘玉[1] 胡娅莉[1] 王亚平[2] 朱海燕[1] 李洁[1] 朱瑞芳[1] 张颖[1] 吴星[1] 杨滢[1] 

机构地区：[1]南京大学医学院附属鼓楼医院妇产科,210008 [2]南京大学医学院附属鼓楼医院医学遗传学实验室,210008

出　　处：《中华妇产科杂志》2008年第11期818-823,共6页Chinese Journal of Obstetrics and Gynecology

基　　金：江苏省医学重点学科项目（XK200709）;江苏省医学领军人才项目（LJ200628）;江苏省社会发展项目（BS2006012）.

摘　　要：目的探讨非多态性位点多重荧光定量PCR（QF-PCR）技术快速产前诊断胎儿染色体非整倍体异常的可行性。方法2006年3月至2007年11月间，收集南京大学医学院附属鼓楼医院早孕期自然流产绒毛组织、中孕期羊水及妊娠晚期的胎儿脐血共63例为研究组。同期60例健康成年人外周血标本（男、女性各30例）作为正常对照组。采用非多态性位点QF-PCR技术检测两组各样本的染色体非整倍体异常情况，以人釉原蛋白基因（AMXY）位点为内参照，根据公式计算剂量系数（DQ）值，DQ值在0．7～1．3之间为正常，〉1．3为染色体扩增，〈0．7则为染色体存在缺失。当2个以上位点电泳峰与AMXY比值异常时，为性染色体数目异常，即各位点与性染色体峰面积比值均〉2．0或〈0．7。将QF-PCR的检测结果与染色体核型分析结果进行比较。结果（1）正常对照组中女性性染色体检测结果为AMX，男性性染色体检测结果为AMX、AMY，与染色体核型分析结果一致。各常染色体DQ值的均值为0．7～1．3，标准差为0．05～0．12。（2）研究组63例中有19例为染色体非整倍体异常，其中13例与核型分析结果一致。与核型分析结果不一致的6例中，1例在18q22．3的CNDP2基因位点表现为三体改变，位于18q12．1的CDH2基因位点表现为正常二倍体，其DQ值为1．28，染色体核型分析为47，XY，＋18；5例非多态性位点QF-PCR结果提示有染色体拷贝数异常，而核型分析结果未见异常，其中1例为47，XY，＋13，核型分析结果为46，XX；1例为Y染色体缺失，而核型分析结果为46，XY；另外3例中1例为47，XX，＋16，2例为47，XX，＋13，而核型分析结果均为46，XX。（3）研究组63例中有44例染色体为正常二倍体，其中36例（82％）与核型分析结果一致。与核型分析结果不一致的8例中，1例为培养失败；2例非多态性QF-PCR结果为正常男性�Objective To explore the feasibility of application of multiplex quantitative fluorescent PCR with non-polymorphic loci in prenatal diagnosis of aneuploidies. Methods From Mar 2006 to Nov 2007, a total of 63 samples were collected from the Department of Obstetrics and Gynecology, Affiliated Drum Tower Hospital of Medical College, Nanjing University, including 54 villous samples obtained for karyotyping because of spontaneous abortion, six amniotic fluid samples of second trimester and three umbilical cord blood samples of third trimester. Blood samples of 60 healthy aduhs were obtained at the same time as a control group, including 30 males and 30 females. Non-polymorphic QF-PCR was performed on both testing group and control group for the detection of aneuploidies. The Amelogenin gene （AMXY） was selected as an internal control, and dosage quotiety （DQ） of each locus was calculated according to the known formula. If DQ was between 0. 7 and 1.3, the sample was considered as normal. If the figure turned out to be 〉 1.3 or 〈 0. 7, a potential duplication or deletion of the corresponding gene or chromosome was indicated. If the results implied numerical abnormalities in more than one euchromosome, sex chromosome aneuploidies should be considered. Cell culture and karyotyping were carried out for every sample simultaneously. The results of non-polymorphic QF-PCR were checked with karyotypes. Results （ 1 ） In the control group, all female samples presented only an AMX peak for sex chromosome while all males showed AMX and AMY amplified peaks. The AMY/AMX ratios were between 0.7 - 1.3, and SD was between 0. 0 -0. 12. （2） Among 19 QF-PCR abnormal cases, 13 cases were proved by karyotyping. Of the six cases which turned out to be conflicting, one case of trisomy 18 shown by karyotyping was not completely detected by QF-PCR, a locus on chromosome 18 implied trisomy, while another turned out to be normal （ DQ = 1.28）. Four cases were detected by non-polymorphic QF-PCR as trisomies but showed normal f

关 键 词：染色体畸变 非整倍性 产前诊断 聚合酶链反应 畸形 

分 类 号：R686[医药卫生—骨科学]