Evaluation of Applied Biosystems' Real-Time Human Quantification Assays




Hybki, Dixie Lee Peters


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To aid the forensic community with its quantification issues, Applied Biosystems is currently developing human specific and human male specific quantification assays using Real-Time PCR (RT-PCR) and TaqMan probes. The human specific assay amplifies an autosomal specific gene, located on chromosome five, while the human male specific assay amplifies a region on the Y chromosome. The purpose of this project was to evaluate the assays with forensic samples to determine if the use of these kits would be appropriate to the forensic community. These kits are not commercially available at the time of this writing. Therefore, several details have been omitted to protect the patent and legal issues that are still pending. It is expected that these assays will surpass the sensitivity and specificity of current methods. This will not only meet, but also exceed the standard set forth by the DAB. By providing additional information such as human male DNA quantification and PCR inhibitor detection, these kits can provide what the forensic community has been lacking. The human male DNA detection and quantification is valuable in providing proof that male DNA was present in an intimate sample from a sexual assault case. This would be especially important in a case in which the offender was a vasectomized male, and for resolving mixtures of the victim and offender’s genetic profiles. The detection of PCR inhibitors for the elimination of futile genetic analysis is a novel component that would provide additional advantages. These kits will offer means for proper quantification to allow for minimal space waste, and allow for successful multiplex PCR within its optimal range. Today, STR analysis will proceed, and is often successful, even if no quantification results are obtained with current methods. The legal system questions this approach. The ability of autosomal specific and Y-chromosome specific RT-PCR quantification assays to assess low level DNA would provide the justification for subsequent analysis that would quiet the legal system’s arguments concerning human quantification.