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Limiting false-positive polymerase chain reaction results: detection of DNA and mRNA to differentiate viable from
Hideo Kobayashi1, Margret Oethinger, Marion J Tuohy
1Pathology and Laboratory Medicine Institute, The Cleveland Clinic Foundation, Cleveland, OH 44195, USA.
Diagnostic Microbiology and Infectious Disease
|July 28, 2009
Abstract:
We compared 2 methods for determining Escherichia coli viability in vitro. A 16S rDNA polymerase chain reaction (PCR) assay detected bacteria irrespective of viability. A groEL mRNA reverse transcriptase PCR was positive for 72 h but later became negative. Detecting mRNA holds promise but is tedious, and groEL may not be the best target.
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02:57Real Time RT-PCR
Real-time reverse transcription-polymerase chain reaction, or Real-time RT-PCR, is an analytical tool used to determine the expression level of target genes. The method involves converting mRNA to complementary DNA with the help of an enzyme known as reverse transcriptase, followed by the PCR amplification of the cDNA. These two processes can be performed simultaneously in a single tube or separately as a two-step reaction.
The real-time quantification of the number of amplified products is...
The real-time quantification of the number of amplified products is...
01:32PCR
Overview