中华医学遗传学杂志
中華醫學遺傳學雜誌
중화의학유전학잡지
CHINESE JOURNAL OF MEDICAL GENETICS
2012年
1期
43-47
,共5页
吴苹%李启杰%夏增亮%张发强%岳琳琳%陈清英%王泓%范春元%夏庆杰
吳蘋%李啟傑%夏增亮%張髮彊%嶽琳琳%陳清英%王泓%範春元%夏慶傑
오평%리계걸%하증량%장발강%악림림%진청영%왕홍%범춘원%하경걸
唐氏综合征%产前诊断%双色竞争性荧光定量聚合酶链反应
唐氏綜閤徵%產前診斷%雙色競爭性熒光定量聚閤酶鏈反應
당씨종합정%산전진단%쌍색경쟁성형광정량취합매련반응
Down syndrome%Prenatal diagnosis%Dual-color competitive quantitative fluorescent polymerase chain reaction
目的 建立一种快速检测唐氏综合征(Down syndrome,DS)的双色竞争性荧光定量聚合酶链反应(dual-color competitive quantitative fluorescent polymerase chain reaction,DCC-QF-PCR)方法,并探讨其应用于DS产前诊断的可能性.方法 提取30例DS患者和60名正常人的外周血DNA,设计DSCR和USC2两基因的特异共用引物和双色特异性TaqMan探针,同一反应管中进行两基因的DCC-QF-PCR,并与DSCR和GAPDH两基因的QF-PCR实验结果进行比较.提取46份羊水胎儿细胞的DNA进行DSCR和USC2两基因的DCC-QF-PCR,并与染色体核型分析结果对比；克隆出DSCR和USC2的单克隆基因片段,定量后进行定量配比的DCC-QF-PCR,并计算DSCR∶USC2拷贝数的比值.结果 DCC-QF-PCR检测中,DS患者DSCR∶USC2拷贝数比值范围为1.41～1.74,显著高于正常人的0.93～1.15;而QF-PCR检测中,DSCR∶GAPDH拷贝数比值范围较DSCR∶USC2增大.46份羊水检测中,3份DSCR∶USC2拷贝数比值分别为1.61、1.64和1.54,为DS患儿,其余43份正常,与染色体核型分析结果一致；DSCR与USC2基因定量配比的DCC-QF-PCR所得DSCR∶USC2拷贝数比值范围与配比值差异无统计学意义(P＞0.05),检测结果较为稳定.结论 DCC-QF-PCR具有准确、快速、需模板量少和费用低廉等特点,该方法在DS的基因诊断和产前基因诊断中有较为广泛的应用前景.
目的 建立一種快速檢測唐氏綜閤徵(Down syndrome,DS)的雙色競爭性熒光定量聚閤酶鏈反應(dual-color competitive quantitative fluorescent polymerase chain reaction,DCC-QF-PCR)方法,併探討其應用于DS產前診斷的可能性.方法 提取30例DS患者和60名正常人的外週血DNA,設計DSCR和USC2兩基因的特異共用引物和雙色特異性TaqMan探針,同一反應管中進行兩基因的DCC-QF-PCR,併與DSCR和GAPDH兩基因的QF-PCR實驗結果進行比較.提取46份羊水胎兒細胞的DNA進行DSCR和USC2兩基因的DCC-QF-PCR,併與染色體覈型分析結果對比；剋隆齣DSCR和USC2的單剋隆基因片段,定量後進行定量配比的DCC-QF-PCR,併計算DSCR∶USC2拷貝數的比值.結果 DCC-QF-PCR檢測中,DS患者DSCR∶USC2拷貝數比值範圍為1.41～1.74,顯著高于正常人的0.93～1.15;而QF-PCR檢測中,DSCR∶GAPDH拷貝數比值範圍較DSCR∶USC2增大.46份羊水檢測中,3份DSCR∶USC2拷貝數比值分彆為1.61、1.64和1.54,為DS患兒,其餘43份正常,與染色體覈型分析結果一緻；DSCR與USC2基因定量配比的DCC-QF-PCR所得DSCR∶USC2拷貝數比值範圍與配比值差異無統計學意義(P＞0.05),檢測結果較為穩定.結論 DCC-QF-PCR具有準確、快速、需模闆量少和費用低廉等特點,該方法在DS的基因診斷和產前基因診斷中有較為廣汎的應用前景.
목적 건립일충쾌속검측당씨종합정(Down syndrome,DS)적쌍색경쟁성형광정량취합매련반응(dual-color competitive quantitative fluorescent polymerase chain reaction,DCC-QF-PCR)방법,병탐토기응용우DS산전진단적가능성.방법 제취30례DS환자화60명정상인적외주혈DNA,설계DSCR화USC2량기인적특이공용인물화쌍색특이성TaqMan탐침,동일반응관중진행량기인적DCC-QF-PCR,병여DSCR화GAPDH량기인적QF-PCR실험결과진행비교.제취46빈양수태인세포적DNA진행DSCR화USC2량기인적DCC-QF-PCR,병여염색체핵형분석결과대비；극륭출DSCR화USC2적단극륭기인편단,정량후진행정량배비적DCC-QF-PCR,병계산DSCR∶USC2고패수적비치.결과 DCC-QF-PCR검측중,DS환자DSCR∶USC2고패수비치범위위1.41～1.74,현저고우정상인적0.93～1.15;이QF-PCR검측중,DSCR∶GAPDH고패수비치범위교DSCR∶USC2증대.46빈양수검측중,3빈DSCR∶USC2고패수비치분별위1.61、1.64화1.54,위DS환인,기여43빈정상,여염색체핵형분석결과일치；DSCR여USC2기인정량배비적DCC-QF-PCR소득DSCR∶USC2고패수비치범위여배비치차이무통계학의의(P＞0.05),검측결과교위은정.결론 DCC-QF-PCR구유준학、쾌속、수모판량소화비용저렴등특점,해방법재DS적기인진단화산전기인진단중유교위엄범적응용전경.
Objective To develop a rapid method for the detection of Down syndrome (DS) using dual-color competitive quantitative fluorescent polymerase chain reaction(DCC-QF-PCR),and to assess its feasibility for the prenatal diagnosis of Down syndrome.Methods DNA was extracted from peripheral blood of 30 DS patients and 60 normal men,common primers for DSCR and USC2 genes and respective TaqMan probes were designed and synthesized.The results of DCC-QF-PCR were compared with those of QF-PCR which measured the ratio between DSCR and GAPDH. Forty-six amniotic fluid samples were assayed with DCC-QF-PCR.The results were compared with that of karyotyping.Monoclone fragments for DSCR and USC2 genes were obtained from direct cloning of PCR products.DCC-QF-PCR was carried out using different DNA ratios of DSCR and USC2 as the template.The dosage ratio between DSCR and USC2 was calculated.Results The gene dosage ratio of the DS patients was 1.41-1.74,which was significantly higher than that of normal men (0.93-1.15).The dosage ratio range of DSCR and GAPDH by QF-PCR was comparatively greater than that of DSCR and USC2.Three samples were diagnosed as DS,which was in good agreement with that of karyotyping analysis.There was no significant difference between the gene dosage ratio from DCC-QF-PCR and that of predetermined(P＞0.05).Conclusion DCC-QF-PCR is an accurate,rapid,and low cost method,which only requires tiny amount of sample and therefore has broad application in the genetic and prenatal diagnosis.
