Co-amplification of the amelogenin and HLA DQ alpha genes: optimization and validation.
An optimized PCR-based system allowing the co-amplification of the HLA DQ alpha and the amelogenin genes has been developed and validated, enabling simultaneous identity testing and sex determination. Sensitivities below 100 pg of human DNA were obtained, using a convenient, high resolution agarose gel system and ethidium bromide staining. Comparison of several co-amplification methods revealed that the best sensitivities and most consistent results were obtained using a hotstart technique employing an inactivating antibody to Taq polymerase. HLA DQ alpha typing results were reliably obtained using the co-amplification process. The sensitivity and ease of this system rendered it directly applicable to forensic analyses. The optimized techniques described here have been validated and successfully applied to forensic cases including People vs. Trujillo, in which the California Superior Court accepted these techniques as scientifically reliable and admissible. Work currently in progress has demonstrated that the described protocol may also be used to co-amplify the amelogenin gene with the AmpliType PM (polymarker) system, allowing identity testing at six loci in addition to sex determination.
Taylor
M S
MS
Technical Associates, Inc., Altadena, CA, USA.
Challed-Spong
A
A
Johnson
E A
EA
D053523:Amelogenin; D001483:Base Sequence; D004247:DNA / Q000032:analysis / Q000235:genetics; D003746:Dental Enamel Proteins / Q000032:analysis / Q000737:chemistry / Q000235:genetics*; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D005554:Forensic Medicine; D005784:Gene Amplification; D006683:HLA-DQ Antigens / Q000032:analysis / Q000737:chemistry / Q000235:genetics*; D006801:Humans; D008297:Male; D016133:Polymerase Chain Reaction; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D012727:Sex Characteristics
D053523:Amelogenin; D003746:Dental Enamel Proteins; D006683:HLA-DQ Antigens; D004247:DNA