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Comparison of STR's Detection Systems for Increased Power of Discrimination and High Through-Put for Case Work SamplesCecelia Crouse, PhD1,Charles Badger2 and Warner Yuen3 Short Tandem Repeat (STR) loci are ubiquitous throughout the human genome. The ultimate goal of STR-based DNA programs is to evaluate specimens for as many repetitive sequences as possible using multiplexed and multicolored STR systems in order to individualize forensic evidence. Over the past two years, the Florida Department of Law Enforcement (FDLE) and the Palm Beach County Sheriff's Office Crime Laboratory (PBSO) have been involved in the development and validation of STRs as an addition to existing forensic DNA programs. The amplification of the CSF1PO/TPOX/TH01 (CTT) multiplex followed by silver staining is currently employed on casework at PBSO. FDLE is investigating fluorescent-based STR technologies using the ABI Prism STR Primer Set/BHO Quadplex provided by Perkin Elmer/Applied Biosystems Division which include the loci F13A1/FES-FPS/vWF/TH01. FDLE is interested in expanding the fluorescent STR repertoire to include CSF1PO and TPOX whereas PBSO is investigating the addition of F13A10/vWF (FFv) for silver staining. Regardless, both laboratories are investigating the same six loci. In order to more fully automate STR technology, the ideal situation would be to multiplex these six and electrophorese the alleles in a single polyacrylamide gel lane. This will not be possible using silver staining as the molecular weights of several of the loci overlap. However, since the loci within the BHO Quadplex are labeled with green (JOE) and blue (FAM) fluorescence and the CTT triplex is labeled with yellow (TAMRA) fluorescence, the ABI Prism 377 DNA Sequencer should be able to analyze the 6 loci in a single lane, thus increasing the power of discrimination as well as incurring high through-put of sample STR information. Further, Promega is developing fluorescent labeled quadriplex systems in which fluorescein-labelled FFFL primers and hex-labeled CTTV primers are used to amplify these STR loci. Using the Hitachi FMBIO fluorescent imaging device, the Promega multiplexed systems can be analyzed in much the same way as the ABI Prism 377 DNA Sequencer. Sexual assault cases are the most common forensic specimens and offer a special challenge of defining mixtures. FDLE and PBSO have evaluated mock sexual assault cases using multiplexed STR loci. Three STR allele detection systems were investigated including silver staining, the ABI Prism 377 DNA Sequencer and the Hitachi FMBIO. The results clearly demonstrate that multiplexed STRs with multicolor-labeling of primers allow for higher throughput whether using the ABI Prism 377 DNA Sequencer or the Hitachi FMBIO and that carry-over of female-to-male or male-to-female fractions is readily discernible. Go to proceedings home page
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