Date of Award

8-1-2007

Degree Type

Restricted Access Professional Report

Degree Name

Master of Science

Field of Study

Cell Biology and Genetics

Department

Graduate School of Biomedical Sciences

First Advisor

Arthur Eisenberg

Second Advisor

Joseph Warren

Third Advisor

John Planz

Abstract

The Maxwell 16 (Promega Corporation, Madison, WI) is a small, self-contained instrument utilizing Promega’s DNA IQ chemistry for the automated extraction of DNA from 16 biological samples simultaneously. Currently, the Maxwell 16 is used in conjunction with the DNA IQ Reference Sample kit for the automated extraction of DNA from forensic and paternity reference samples. Promega Corporation is currently in the development of the DNA IQ Casework Sample kit with the intent of using the Maxwell 16 instrument in the extraction of DNA from forensic evidentiary samples. Modifications have been made to Maxwell 16 to allow the elution of DNA in a smaller volume (Low-Elution Volume (LEV) configuration) that when used in conjunction with the DNA IQ Casework Sample kit would optimize DNA yield from forensic casework samples. However, the limited quantity and the low quality of forensic casework samples are significant challenges facing most automated DNA extraction systems. An evaluation study was conducted to test the performance of the Maxwell 16 instrument along with the DNA IQ Casework Sample kit for processing forensic evidentiary samples. Mock evidentiary items used for the evaluation consisted of blood, semen, tissue, and touch samples that are routinely encountered in forensic casework. Prior to loading the Maxwell 16 instrument samples were first digested using the Tissue and Hair Extraction kit (Promega Corporation) designed for optimum DNA recovery. The extraction of DNA from sexual assault samples typically requires the separation of the sperm DNA deposited by the assailant from the vaginal epithelial cell DNA from the victim. Promega Corporation has developed the Differex System which utilizes a manual phase separation technique to obtain both a sperm fraction and an epithelial cell fraction. After separation and digestion, samples were then added to the DNA IQ Casework Sample kit cartridges for automated DNA purification using the Maxwell 16. For comparison, DNA was also obtained from replicate samples processed with the Differex System following the standard organic extraction method. The evaluation of the Differex system with mock sexual assault samples was conducted in order to determine if this differential extraction process can be used in conjunction with Maxwell 16 to improve case processing efficiency. To evaluate the performance of the Maxwell 16 and the DNA IQ Casework Sample kit, replicate samples were prepared and processed using UNTHSC’s standard organic extraction methodology. The DNA obtained by both methodologies was quantified using the Applied Biosystems (AB) Quantifiler Human DNA Quantification kit (Foster City, CA) and the AB 7500 Real-Time PCR system and then amplified using the PowerPlex 16 kit (Promega Corporation). The amplified DNA was analyzed using the AB 3130xl Genetic Analyzer and the resulting STR electropherograms were analyzed to obtain profiles using GeneMapper ID v3.2 (Applied Biosystems). In addition to assessing the quantity and the quality of the DNA obtained, blank cartridges were simultaneous processed with the mock forensic samples to demonstrate whether the Maxwell 16 could introduce cross-contamination between samples. The overall performance and the cost-effectiveness are the ultimate criteria to help determine the utility of the Maxwell 16 in the processing of forensic evidentiary casework samples.

Comments

W 4.8 A415E 2007

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