Validation of internal controls for extraction and amplification of nucleic acids from enteric viruses in water samples.

ABSTRACT: Inhibitors that reduce viral nucleic acid extraction efficiency and interfere with cDNA synthesis and/or polymerase activity affect the molecular detection of viruses in aquatic environments. To overcome these significant problems, we developed a methodology for assessing nucleic acid yields and DNA amplification efficiencies for environmental water samples. This involved adding particles of adenovirus type 5 and murine norovirus and newly developed primer-sharing controls, which are amplified with the same primer pairs and result in the same amplicon sizes as the targets, to these samples. We found that nucleic acid loss during the extraction process, rather than reverse transcription-PCR (RT-PCR) inhibition, more significantly attributed to underestimation of the presence of viral genome

SUBMITTER: Hata A 

PROVIDER: S-EPMC3127680 | biostudies-literature | 2011 Jul

REPOSITORIES: biostudies-literature