P. Sean Walsh, Nicola Fildes and Kathy Lazaruk
Perkin-Elmer, Applied Biosystems Division, Foster City, CA
ABSTRACT
Short Tandem Repeat (STR) loci are very informative forensic markers because many of the loci exhibit a high degree of polymorphism in human populations. Tetranucleotide repeat loci are abundant in the human genome and are amenable to analysis by the Polymerase Chain Reaction (PCR). This report describes the general approach we have undertaken for the development and validation of STR kits that provide reagents for co-amplification of highly informative STR loci and detailed protocols for fluorescence detection. Three STR loci are co-amplified using priers that have been labeled with the same color fluorescent dye. The PCR amplification has been optimized to provide specific, robust amplification of forensic samples, resulting in balanced signals across each of the loci. The fluorescently tagged STR alleles are then resolved and detected by gel electrophoresis on the ABI PRISM 373 or 377 DNA Sequencer or by capillary electrophoresis on the ABI PRISM 310 Genetic Analyzer. These instruments provide multiple dye color detection, automatic allele peak quantitation and automatic allele size determination using an in-lane standard. Three STR triplex kits are currently under development. Each STR triplex will be labeled with a different color fluorescent dye. Therefore, three PCR reactions from a single sample may be combined with an internal lane standard to type nine STR loci in a single lane. Each STR kit will be validated according to TWGDAM guidelines.
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