Semester

Summer

Date of Graduation

2004

Document Type

Dissertation

Degree Type

PhD

College

School of Medicine

Department

Microbiology, Immunology, and Cell Biology

Committee Chair

Clifton P. Bishop.

Abstract

DNA analysis by Polymerase Chain Reaction (PCR) allows for the unambiguous identification of the person from whom a biological sample was derived but provides no information about when the sample was deposited. The ability to determine the age of blood or a biological sample would be of significant benefit to the forensic science community. This would provide a temporal linkage between the blood of the perpetuator and the commission of a crime or it could be used to exclude evidence that does not correspond to the time a crime was committed. It was hypothesized that ex vivo RNA decay was a predictable process and real-time PCR indicated that different types of RNA and different sized regions of RNA influenced our capabilities of RNA detection in dried blood stains. The ratio/difference in Ct values between different types and regions of RNA (rRNA vs. mRNA), was identified to change over time in blood stains. Under the conditions examined in the present study, the ratio between the identified RNA species changed in a consistent and reliable way. Environmental influences on RNA detection in dried blood were explored; temperature and humidity both influenced RNA detection but full spectrum light did not. In order to establish a shorter time frame of detection, DNA microarrays were used to identify RNAs that may undergo changes in RNA levels ex vivo due to a stress response. Preliminary results identified several candidate RNA that had up-regulated levels from 4-96 fold in the first 1-4 hours after deposition. Although other approaches have been used in the past to estimate the age of a biological sample, this approach offers the following advantages: DNA and RNA can be co-isolated from the same sample; probes can be made species specific; relatively small samples can be assayed; and, the window of time estimates for dried blood is at least 0 to 150 days. This work provides a starting point for a technique that may prove useful in estimating the age of a biological sample. With the integration of the above mentioned techniques, this method has the potential to solve a problem the forensic community has been investigating for over 100 years.

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