QUANTITATIVE POLYMERASE CHAIN-REACTION BY MONITORING ENZYMATIC-ACTIVITY OF DNA-POLYMERASE

被引:21
作者
YANG, BZ [1 ]
YOLKEN, R [1 ]
VISCIDI, R [1 ]
机构
[1] JOHNS HOPKINS UNIV,SCH HYG & PUBL HLTH,DEPT IMMUNOL & INFECT DIS,BALTIMORE,MD 21205
关键词
D O I
10.1006/abio.1993.1014
中图分类号
Q5 [生物化学];
学科分类号
071010 ; 081704 ;
摘要
Nucleic acid amplification by polymerase chain reaction (PCR) is a very powerful technique in terms of sensitivity but is limited in terms of ability to perform accurate quantitation. While there is a theoretical correlation between copies of input target sequence and those of PCR product, the quantitative nature of this relationship is obscured by unpredictable variations in reaction conditions and by inhibitory and/or stimulatory substances which might be present in sample preparations, especially those derived from biological fluids. To reliably estimate copies of input DNA target from PCR product, we designed a combination of internal and external control systems coupled to DNA/RNA hybridization and enzymatic immunodetection techniques. The internal control system served to monitor amplification efficiency and to correct for the effects of inhibitors or stimuli on the efficiency of the DNA amplification. The assay is quantitative, nonisotopic, and can be widely applied to assessment of the quantity of DNA present in a wide range of preparations. © 1993 Academic Press, Inc.
引用
收藏
页码:110 / 116
页数:7
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