Date of Award

8-1-2006

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

Joseph Warren

Second Advisor

John Planz

Third Advisor

Arthur Eisenberg

Abstract

The introduction of a new instrument into an accredited laboratory requires a documented internal validation. Validations typically include sensitivity, precision, and mixture studies. These tests assess the reliability and efficiency of the instrument and allow for interpretation guidelines to be established. This project consisted of a validation of Applied Biosystems’ 3130xl Genetic Analyzer and an evaluation of three mitochondrial DNA amplification primer sets for the control region. The validation was designed to evaluate the efficacy, robustness, and working limitations of the 3130xl instrument by performing a sensitivity study. A sensitivity study was performed using DNA sample dilutions, which were quantified using ABIs Quantifiler system to measure the amount of total nuclear DNA content in the samples. The samples were amplified on the GeneAmp PCR System 9700 in triplicate to evaluate stochastic activity. Three different primer sets were utilized which allow for the amplification of different regions of the human mitochondrial genome control region. After amplification, the quality and quantity of the DNA in all the samples was assessed using the Agilent 2100 BioAnalyzer, and subsequent sequence analysis was performed on the 3130xl Genetic Analyzer. Preliminary work has begun on a mixture study, but due to lack of time and reagents, this study was not completed and will have to be performed at a later date. All sequence data from the sensitivity study was evaluated using Sequencher version 4.1.4Fb19

Comments

W 4.8 D249V 2006

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