Synergy between DNA polymerases increases polymerase chain reaction inhibitor tolerance in forensic DNA analysis.
The success rate of diagnostic polymerase chain reaction (PCR) analysis is lowered by inhibitory substances present in the samples. Recently, we showed that tolerance to PCR inhibitors in crime scene saliva stains can be improved by replacing the standard DNA polymerase AmpliTaq Gold with alternative DNA polymerase-buffer systems (Hedman et al., BioTechniques 47 (2009) 951-958). Here we show that blending inhibitor-resistant DNA polymerase-buffer systems further increases the success rate of PCR for various types of real crime scene samples showing inhibition. For 34 of 42 "inhibited" crime scene stains, the DNA profile quality was significantly improved using a DNA polymerase blend of ExTaq Hot Start and PicoMaxx High Fidelity compared with AmpliTaq Gold. The significance of the results was confirmed by analysis of variance. The blend performed as well as, or better than, the alternative DNA polymerases used separately for all tested sample types. When used separately, the performance of the DNA polymerases varied depending on the nature of the sample. The superiority of the blend is discussed in terms of complementary effects and synergy between the DNA polymerase-buffer systems.
Hedman
Johannes
J
Department of Applied Microbiology, Lund University, SE-221 00 Lund, Sweden.
Nordgaard
Anders
A
Dufva
Charlotte
C
Rasmusson
Birgitta
B
Ansell
Ricky
R
Rådström
Peter
P
D002021:Buffers; D004247:DNA / Q000032:analysis*; D004259:DNA-Directed DNA Polymerase / Q000737:chemistry / Q000378:metabolism*; D044707:Forensic Sciences; D016133:Polymerase Chain Reaction / Q000379:methods*; D012463:Saliva / Q000737:chemistry
D002021:Buffers; D004247:DNA; D004259:DNA-Directed DNA Polymerase