Exogenous contaminating DNA in Taq polymerases: A method to avoid false-positive results when detecting the blaTEM gene

J Microbiol Methods. 2019 May:160:36-41. doi: 10.1016/j.mimet.2019.03.018. Epub 2019 Mar 20.

Abstract

In the course of developing an assay to identify genes responsible for antibiotic resistance in gram-negative bacteria, it has been found that standard (not DNA-free) Taq DNA polymerases were contaminated with blaTEM gene fragments that varied in length and quantities. The complete blaTEM gene sequence was either absent or was detected in infinitesimal amounts. We developed an approach to avoid false-positive findings caused by contaminating blaTEM gene sequences in conventional polymerases. The method is based on selection of a target sequence to be detected within the blaTEM gene in such a way that the chosen sequence is amplified with primers incapable of amplifying contaminating DNA sequences of the polymerase.

Keywords: Contaminating DNA; False-positive results; Microarray; Taq DNA polymerase; bla(TEM) gene.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • DNA Contamination*
  • DNA Primers / chemistry
  • DNA, Bacterial / genetics*
  • Escherichia coli / genetics
  • False Positive Reactions
  • Polymerase Chain Reaction / methods*
  • Taq Polymerase / analysis*

Substances

  • DNA Primers
  • DNA, Bacterial
  • Taq Polymerase