Chang D, Wang M, Ou W C, Tsai R T, Fung C Y, Hwang Y J
Department of Medicine, Chung Shan Medical and Dental College, Taichung, Taiwan, Republic of China.
J Virol Methods. 1996 Apr 26;58(1-2):131-6. doi: 10.1016/0166-0934(95)02001-2.
In order to develop a simple and sensitive method for detecting human polyomavirus DNA in the urine of patients by the polymerase chain reaction (PCR), it was found that the viral DNA could be released from urine by proteinase K and then amplified by PCR directly, without additional treatment such as ultracentrifugation or DNA extraction. Direct PCR amplification of viral DNA from urine was volume limited and 5 microliters of urine appeared to be the optimum amount for direct PCR amplification. When the urine volume was greater than 10 microliters, the results of PCR were inconsistent. However, the urine volume could be increased after dialysis to remove possible inhibitor(s) which may interfere with PCR. Direct PCR amplification of patient urine is convenient and eliminates several steps which can cause loss of DNA from the sample.
为了开发一种通过聚合酶链反应（PCR）检测患者尿液中人类多瘤病毒DNA的简单且灵敏的方法，研究发现蛋白酶K可从尿液中释放病毒DNA，然后直接通过PCR进行扩增，无需超速离心或DNA提取等额外处理。从尿液中直接进行病毒DNA的PCR扩增受体积限制，5微升尿液似乎是直接PCR扩增的最佳量。当尿量大于10微升时，PCR结果不一致。然而，透析后可增加尿量以去除可能干扰PCR的抑制剂。对患者尿液进行直接PCR扩增很方便，且省去了几个可能导致样本DNA丢失的步骤。
