Forensic Stain Identification By RT-PCR Analysis (Updated)

• NCJ Number: 236537
• Author(s): Trisha Conti, Ph.D.; Eric Buel, Ph.D.
• Date Published: 2011
• Length: 88 pages
• Grant Number(s): 2007-DN-BX-K149
• Agencies: NIJ-Sponsored
• Publication Type: Report (Study/Research)

Annotation

This report describes the methodology and findings of a project that developed a RNA/DNA co-isolation technique that extracts both nucleic acids of sufficient quality and quantity for downstream real-time PCR and STR analyses.

Abstract

The project's first goal was to identify the best method to co-extract DNA and RNA from a variety of stain types. By using one extraction step, a DNA sample would be ready and waiting for STR profiling if the RNA screening assay deemed it worthy of such analysis. In addition, obtaining RNA and DNA from a single stain would present the possibility of conclusions being drawn regarding the identity of one stain that may hot hold true for a nearby stain. Based on preliminary experiments, the TRIzol method was identified as an efficient and straight-forward procedure for the isolation of DNA and RNA. Still, there are several disadvantages which caused researchers to seek an alternative isolation procedure. A homebrew extraction method was developed that worked for RNA and DNA. It is faster than the TRIzol method and produces significantly better DNA yields. The project then identified gene candidates that were specific for each tissue of interest. The candidates used throughout the project were identified through literature review, Gene, and other databases. Researchers tested the specificity and sensitivity of each assay by analysis of mRNA isolated from the body fluid of interest (seminal fluid) or control RNA (brain). The diverse sample bank was used to assess the specificity of the candidate tissue-specific genes. These samples included blood, semen, saliva, menstrual blood vaginal secretions, kidney, colon, adipose, skin and control human RNAs (brain, heart, liver, kidney, and intestine). The completion of project components produced the conclusion that DNA and RNA can be co-extracted and the RNA fraction used in multiplexed real-time PCR assays. 3 figures and 39 references