Multiplex Amplification and Automated Fluorescent Typing of Short Tandem Repeat (STR) Loci: The French Experience

Abstract

The great majority of the biological samples available in forensic investigations for DNA identification purposes are severely degraded or present in minute amounts and can not be analysed by conventional RFLP (Restriction Fragment Length Polymorphism) methods using single locus probing. The polymerase Chain Reaction (PCR) amplification of short tandem repeat (STR) loci appears to be an efficient, sensitive and rapid DNA identification system for highly degraded samples. In order to use STR-PCR for forensic purposes or parental testing in routine analyses, a significant number of individuals (>100) has to be typed to determine accurately the allelic frequencies in the population of interest. Allele frequencies for the HUMTH01 (TH01) and HUMFESFPS (FES) STR loci were obtained through singleplex PCR and manual silver staining detection. The purchase of the automated ABI 373A sequencer allowed us to establish the allelic distributions for HUMVWA31A (VWA) and HUMF13A1 (F13) STR loci by duplex amplification and automated fluorescence-based detection. The quadruplex PCR of these four STR loci was then developped and performed on biological samples of various origins and appear to be robust. Furthermore, we studied a pentaplex-PCR, including 4 STR loci (D6S502, D18S51, D21S11 and HUMFIBRA) and the X-Y homologous gene amelogenin for sex determination: allelic frequencies were also determined.