title: Multiplex short tandem repeat amplification of low template DNA samples with the addition of proofreading enzymes.
abstract: With <100 pg of template DNA, routine short tandem repeat (STR) analysis often fails, resulting in no or partial profiles and increased stochastic effects. To overcome this, some have investigated preamplification methods that include the addition of proofreading enzymes to the PCR cocktail. This project sought to determine whether adding proofreading polymerases directly in the STR amplification mixture would improve the reaction when little template DNA is available. Platinum Taq High Fidelity and GeneAmp High Fidelity were tested in Profiler Plus™ STR reactions alone and in combination with AmpliTaq(®) Gold. All reactions included the additional step of a post-PCR purification step. With both pristine low template DNA and casework samples, the addition of these polymerases resulted in comparable or no improvement in the STR amplification signal. Further, stochastic effects and artifacts were observed equally across all enzyme conditions. Based on these studies, the addition of these proofreading enzymes to a multiplex STR amplification is not recommended for low template DNA work.
authors.affiliation: Department of Forensic Science and Biology, Virginia Commonwealth University, 1000 West Cary Street, PO Box 842012, Richmond, VA 23284, USA.
authors.forename: Carey P
authors.initials: CP
authors.lastname: Davis
authors.forename: Lynzee A
authors.initials: LA
authors.lastname: Chelland
authors.forename: Victoria R
authors.initials: VR
authors.lastname: Pavlova
authors.forename: María J
authors.initials: MJ
authors.lastname: Illescas
authors.forename: Kelly L
authors.initials: KL
authors.lastname: Brown
authors.forename: Tracey Dawson
authors.initials: TD
authors.lastname: Cruz
mesh_terms: D000483:Alleles; D001769:Blood / Q000378:metabolism; D001842:Bone and Bones / Q000378:metabolism; D004247:DNA / Q000032:analysis* / Q000235:genetics; D016172:DNA Fingerprinting / Q000379:methods*; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism*; D003878:Dermatoglyphics; D019075:Electrophoresis, Capillary; D006801:Humans; D007180:Incisor / Q000378:metabolism; D016133:Polymerase Chain Reaction; D013232:Steel; D020080:Tandem Repeat Sequences
chemical_list: D013232:Steel; D004247:DNA; D004259:DNA-Directed DNA Polymerase