title: Residual DNA in thermostable DNA polymerases - a cause of irritation in diagnostic PCR and microarray assays.
abstract: In a validation trial of a DNA microarray test for chlamydiae we repeatedly observed false-positive PCR amplicons from truly negative samples and non-template controls. Various PCR tests, microarray hybridization and DNA sequencing, revealed that residual Escherichia coli DNA from thermostable DNA polymerases was the cause of this cross-reaction. A subsequent survey showed that only five out of 23 commercial polymerases were free of E. coli DNA. When designing generic oligonucleotide sequences for PCR and PCR microarray-based assays one should be aware of such possible internal contamination, particularly when the target organism is E. coli.
authors.affiliation: CLONDIAG chip technologies GmbH, Loebstedter Strasse 103-105, 07749 Jena, Germany. ralf@clondiag.com
authors.forename: Ralf
authors.initials: R
authors.lastname: Ehricht
authors.forename: Helmut
authors.initials: H
authors.lastname: Hotzel
authors.forename: Konrad
authors.initials: K
authors.lastname: Sachse
authors.forename: Peter
authors.initials: P
authors.lastname: Slickers
mesh_terms: D002693:Chlamydiaceae / Q000235:genetics / Q000302:isolation & purification; D004269:DNA, Bacterial / Q000032:analysis* / Q000302:isolation & purification; D004259:DNA-Directed DNA Polymerase / Q000737:chemistry* / Q000378:metabolism; D004866:Equipment Contamination; D004926:Escherichia coli / Q000235:genetics / Q000302:isolation & purification; D046228:Microarray Analysis / Q000592:standards*; D016133:Polymerase Chain Reaction / Q000592:standards*; D011933:Reagent Kits, Diagnostic / Q000382:microbiology*; D012680:Sensitivity and Specificity; D019914:Taq Polymerase / Q000737:chemistry; D013696:Temperature
chemical_list: D004269:DNA, Bacterial; D011933:Reagent Kits, Diagnostic; D019914:Taq Polymerase; D004259:DNA-Directed DNA Polymerase