Polymerase chain reaction
Nuclear secrets: Quality control of forensic samples using qPCR
Real-time quantitative PCR (qPCR) is emerging as the new method of choice for analysing nuclear DNA levels in forensic samples. A useful addition to this technique would be the ability to quantitatively assess the quality of this DNA, as forensic samples often present considerable DNA degradation that must be taken into account during subsequent genotyping procedures.
Until recently, slot blot hybridisations were the most common method used by DNA analysts to evaluate forensic samples prior to genotyping. However, in comparison with the increasingly sophisticated genotyping methods (which usually involve the analysis of short tandem repeats, or STRs), slot blot hybridisations are less sensitive and less accurate, and reveal nothing about the quality of the DNA. They are also relatively time-consuming and labour-intensive, and do not lend themselves readily to automation.
Given the need of forensic laboratories to efficiently process increasing numbers of DNA samples and the fact that many of these of poor quality, there is growing incentive to improve the success rate of first-pass genotyping. This rate could be improved by knowing more about the quantity and quality of the DNA prior to STR analysis. It comes as no wonder, then, that forensic scientists are turning to qPCR for a quicker and more precise measurement of nuclear DNA amounts.
Recently, a team of investigators around Katie Swango, of the Jan Bashinski DNA Laboratory of the California Department of Justice, has taken this approach one step further. In an advance online publication in Forensic Science International, the California group describes in detail the development and experimental validation of a multiplex qPCR assay to simultaneously test both the quantity and quality of nuclear DNA in forensic samples. By amplifying two nuclear DNA sequences of different lengths, both relevant to STR amplification targets, the degree of sample degradation could readily be quantified.
This rigorously tested protocol should prove useful for forensic scientists wishing to improve the throughput of their sample processing and genotyping platforms.
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