Reduced volume PCR amplification reactions using the AmpFlSTR Profiler Plus kit.
The forensic community continues to seek improvements in DNA typing methods on aspects such as sensitivity and efficacy. Reducing the volume of the AmpFlSTR Profiler Plus reagents offered greater sensitivity and improved the chance of obtaining useful results for samples with very low quantities of DNA and multiple source samples. On the downside, amplifications initiated with less than 0.4 ng of DNA exhibited a twofold increase in the standard deviation of peak ratios. This research suggested a twofold approach to analyzing samples. For samples with greater than 0.25 ng of DNA, a 25 microL reaction is appropriate. Samples that did not demonstrate quantifiable results, or that have less than 0.25 ng, can be amplified by drying the sample directly in the PCR tube and amplifying in a 5 microL reaction. The analyst can expect at least limited results with as little as 0.03 ng of DNA in the 5 microL reaction.
Gaines
Michelle L
ML
North Louisiana Criminalistics Laboratory, Shreveport 71101, USA.
Wojtkiewicz
Patrick W
PW
Valentine
Jennifer A
JA
Brown
Connie L
CL
D000483:Alleles; D016172:DNA Fingerprinting / Q000379:methods*; D004586:Electrophoresis / Q000379:methods; D005554:Forensic Medicine / Q000379:methods; D006801:Humans; D016133:Polymerase Chain Reaction / Q000295:instrumentation* / Q000379:methods*; D012680:Sensitivity and Specificity; D013223:Statistics as Topic / Q000379:methods