检测AML多药耐药基因mRNA表达的多重半巢式逆转录PCR方法的建立

Detection of MDR gene mRNA expression in AML by using multiplex reverse transcription-PCR

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 multiplex reverse transcription-PCR detetion system to test multidrug resistance （MDR） gene mRNA expression in acute myelocytic leukemia（AML）. Methods Five pairs of primes specific to five MDR gene and one pair to b-MG gene were designed and the single-tube six-complex reverse transcription-PCR system was established. Results Five MDR gene could be detected simultaneously in the multiplex PCR system, The result of multiplex PCR were according with that of one single PCR in the detection of clinical specimens. Conclusion This method is sensitive, specific, and has advantages of speed in performance when used to dectct the MDR gene mRNA expression in HL60/VCR cell line and bone marrows of AML patients, It may also be applied to process a batch of sample simultaneously in one setting.