Molecular assay for screening and quantifying DNA in biological evidence: the modified Q-TAT assay.
A method is described for the quantitation of total human and male DNA. Q-TAT utilizes end-point, multiplex polymerase chain reaction (PCR) amplification of the amelogenin and SRY loci to quantify DNA and incorporates a cloned nonhuman template to detect PCR inhibition. Standard curves of fluorescence from amelogenin or SRY amplicons were generated from amplification of known amounts of NIST traceable SRM-female or SRM-male DNA. Curves showed good linearity up to 500 pg of SRM-template (R(2) > 0.99) and reliably estimated total and male DNA content in casework samples. The nonhuman pRL(null) template included in each PCR was a sensitive indicator of known PCR inhibitors including EDTA, hemin, blue denim dye, and humic acid. Finally, the SRY amplicon was a sensitive indicator of male DNA and, in mixtures, could reliably estimate male DNA present in an excess of female DNA. The Q-TAT multiplex is a reliable quantitation method for forensic DNA typing.
Wilson
Jon
J
Police Laboratory, Tulsa Police Department, Tulsa, OK, USA.
Fuller
Valerie
V
Benson
Gifty
G
Juroske
Denise
D
Duvall
Eric
E
Fu
Jun
J
Pritchard
Jane
J
Allen
Robert W
RW
D041322:Chromosomes, Human, Y; D004396:Coloring Agents; D004247:DNA / Q000302:isolation & purification; D016172:DNA Fingerprinting / Q000379:methods*; D017931:DNA Primers; D003746:Dental Enamel Proteins / Q000235:genetics; D004492:Edetic Acid; D005260:Female; D024282:Genes, sry / Q000235:genetics*; D006427:Hemin; D006801:Humans; D006812:Humic Substances; D049410:Luciferases, Renilla / Q000235:genetics*; D008297:Male; D010957:Plasmids / Q000235:genetics; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000378:metabolism; D017422:Sequence Analysis, DNA; D013698:Templates, Genetic
D004396:Coloring Agents; D017931:DNA Primers; D003746:Dental Enamel Proteins; D006812:Humic Substances; D012333:RNA, Messenger; D006427:Hemin; D004247:DNA; D004492:Edetic Acid; D049410:Luciferases, Renilla