一步多重PCR同时检测甲型肝炎和脊髓灰质炎病毒研究

Simultaneous detection of both hepatitis A virus and poliovirus by a one-step multiplex polymerase chain reaction

建立的一步PCR方法即反转录和PCR在同一管中进行 ,同时检测甲型肝炎和脊髓灰质炎病毒病毒RNA。实验中对不同的反转录温度以及一步多重PCR的特异性和灵敏度进行了探讨。结果表明 :42℃、5 0℃反转录时polio有非特异性条带出现 ,6 0℃反转录特异性较好 ,而HAV在三种不同的反转录温度下均得到特异性较好的条带 ;应用一步PCR同时检测两种病毒所得到的条带分别与检测单一病毒所得条带一致 ,半套式PCR也得到了预期的条带 ;应用一步PCR同时检测两种病毒与检测单一病毒的灵敏度基本一致 ,但在同等反应条件下后者的反应效率高于前者 ,特别是在检测HAV时。

An one step procedure was developed using the PCR(polymerase chain reaction)for the detection of both hepatitis A virus and poliovirus RNA,in which reverse transcription and PCR were processed in one tube.Different reverse transcription temperatures, the specility and sensitivity of one step multiplex PCR were studied in experiment.The results indicated that poliovirus had non specific bands when reverse transcription was carried out at 42℃ and 50℃,but only specific hand appeared at 60℃, As far as HAV, the specific bands were achieved at the three temperatures.The bands of one species of virus were in correlation with that of two species of virus when detected by one step PCR.The expected bands were also achieved in the semi nested PCR.The sensitivity of two species of virus detected by one step PCR was nearly parallel to that of one species of virus,but the reaction efficiency of the latter was higher than the former at the same reaction condition,especially to HAV.