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目的建立多重逆转录和半巢式PCR的方法检测急性髓性白血病(AML)多药耐药(MDR)基因mRNA表达。方法分别设计针对5种MDR基因和1种内参照基因的特异性引物,采用多重逆转录和半巢式PCR检测耐药细胞株HL60/VCR和难治性AML患者骨髓标本中的MDR基因mRNA表达。结果应用此多重逆转录和半巢式PCR反应系统,可以特异的在一个反应管内同时检测到5种耐药基因。结论该方法对于检测AML患者MDR基因mRNA表达具有快速、灵敏、简便等特点,能快速同时分析大量样本,对MDR研究、诊断和治疗有一定应用前景。
Objective To establish a multiplex reverse transcription and semi-nested PCR method for the detection of multidrug resistance (MDR) mRNA in acute myeloid leukemia (AML). Methods Specific primers targeting five MDR genes and one internal reference gene were designed and used to detect MDR mRNA expression in bone marrow samples of drug resistant HL60 / VCR and refractory AML patients by multiplex reverse transcription and semi-nested PCR . Results Using this multiplex reverse transcription and semi-nested PCR reaction system, we could detect 5 kinds of drug resistance genes in one reaction tube at the same time. Conclusion The method is rapid, sensitive and simple for the detection of MDR gene mRNA expression in AML patients. It can rapidly and simultaneously analyze a large number of samples and has certain application prospect for MDR research, diagnosis and treatment.