中华消化杂志
中華消化雜誌
중화소화잡지
Chinese Journal of Digestion
2012年
7期
466-468
,共3页
聚合酶链反应%多态性,限制性片段长度%DNA甲基化%粪便%胰腺肿瘤%脑啡肽类%基因,K-ras%突变
聚閤酶鏈反應%多態性,限製性片段長度%DNA甲基化%糞便%胰腺腫瘤%腦啡肽類%基因,K-ras%突變
취합매련반응%다태성,한제성편단장도%DNA갑기화%분편%이선종류%뇌배태류%기인,K-ras%돌변
Polymerase chain reaction%Polymorphism,restriction fragment length%DNA methylation%Feces%Pancreatic neoplasms%Enkephalins%Genes,K-ras%Mutation
目的 评价甲基化特异性聚合酶链反应(MSP)检测粪便ppENK基因高甲基化用于诊断胰腺癌的可行性.方法 收集胰腺癌患者24例和对照6例的新鲜粪便标本.采用MSP检测全部粪便标本中ppENK的甲基化状态；采用PCR检测全部粪便标本中野生型ppENK的阳性率.采用PCR-限制性片段长度多态性(RFLP)检测粪便K-ras的突变情况.将胰腺癌细胞PC3单细胞悬液加入同一健康者粪便标本,MSP法检测ppENK甲基化阳性,计算甲基化阳性时需掺入的胰腺癌细胞的最少数量.结果 30份粪便标本的甲基化检出率为0(0/30),非甲基化检出率为10％(3/30),野生型ppENK检出率为6.7％ (2/30)；PCR-RFLP可检测出所选10份野生型ppENK阴性的胰腺癌标本中的8份,其中7份有K- ras第12位密码子突变.MSP方法能够检测到粪便中ppENK甲基化条带所需胰腺癌细胞的数量至少为50个/ml.结论 采用MSP方法检测胰腺癌患者粪便标本的ppENK基因甲基化状态,尚不能成为筛查和诊断胰腺癌的方法.
目的 評價甲基化特異性聚閤酶鏈反應(MSP)檢測糞便ppENK基因高甲基化用于診斷胰腺癌的可行性.方法 收集胰腺癌患者24例和對照6例的新鮮糞便標本.採用MSP檢測全部糞便標本中ppENK的甲基化狀態；採用PCR檢測全部糞便標本中野生型ppENK的暘性率.採用PCR-限製性片段長度多態性(RFLP)檢測糞便K-ras的突變情況.將胰腺癌細胞PC3單細胞懸液加入同一健康者糞便標本,MSP法檢測ppENK甲基化暘性,計算甲基化暘性時需摻入的胰腺癌細胞的最少數量.結果 30份糞便標本的甲基化檢齣率為0(0/30),非甲基化檢齣率為10％(3/30),野生型ppENK檢齣率為6.7％ (2/30)；PCR-RFLP可檢測齣所選10份野生型ppENK陰性的胰腺癌標本中的8份,其中7份有K- ras第12位密碼子突變.MSP方法能夠檢測到糞便中ppENK甲基化條帶所需胰腺癌細胞的數量至少為50箇/ml.結論 採用MSP方法檢測胰腺癌患者糞便標本的ppENK基因甲基化狀態,尚不能成為篩查和診斷胰腺癌的方法.
목적 평개갑기화특이성취합매련반응(MSP)검측분편ppENK기인고갑기화용우진단이선암적가행성.방법 수집이선암환자24례화대조6례적신선분편표본.채용MSP검측전부분편표본중ppENK적갑기화상태；채용PCR검측전부분편표본중야생형ppENK적양성솔.채용PCR-한제성편단장도다태성(RFLP)검측분편K-ras적돌변정황.장이선암세포PC3단세포현액가입동일건강자분편표본,MSP법검측ppENK갑기화양성,계산갑기화양성시수참입적이선암세포적최소수량.결과 30빈분편표본적갑기화검출솔위0(0/30),비갑기화검출솔위10％(3/30),야생형ppENK검출솔위6.7％ (2/30)；PCR-RFLP가검측출소선10빈야생형ppENK음성적이선암표본중적8빈,기중7빈유K- ras제12위밀마자돌변.MSP방법능구검측도분편중ppENK갑기화조대소수이선암세포적수량지소위50개/ml.결론 채용MSP방법검측이선암환자분편표본적ppENK기인갑기화상태,상불능성위사사화진단이선암적방법.
Objective To evaluate the feasibility of testing the high methylation of ppENK gene in stool with methylation-specific polymerase chain reaction (MSP) assay in pancreatic cancer diagnosis.Methods Twenty-four fresh stool samples of pancreatic cancer patients and six healthy control samples were collected.The methylation status of ppENK gene in all the stool samples was detected by MSP assay.The positive rate of wild-type ppENK gene in all the stool samples was determined by polymerase chain reaction (PCR).And 10 non wild-type ppENK gene negative pancreatic cancer samples were collected,and K-ras gene mutation was detected by PCR-restriction fragment length polymorphism (RFLP).The single cell suspension of pancreatic cancer PC3 cell line was added into stool sample from the same healthy individual,the positive rate of ppENK gene methylation was detected by MSP assay.The minimum number of pancreatic cancer cell was calculated when methylation was positive.Results The rate of methylation detection in 30 samples was 0 (0/30); and the rate of non-methylation detection was 10％ (3/30).The rate of wild-type ppENK detection was 6.7％ (2/30).By PCR-RFLP assay,eight were successfully amplified and seven had mutation in 12th code of K-ras gene in 10 selected wild-type ppENK gene negative pancreatic cancer samples.The minimum number of pancreatic cancer cells needed for ppENK methylation band positive detected by MSP was 50 cell/ml.Conclusion Detecting ppENK gene methylation status in stool samples of pancreatic cancer patients by MSP assay has not yet become the method of pancreatic cancer screening and diagnosis.
