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Mitochondrial DNA

Mitochondrial DNA. Observations on Frye/Daubert Issues Associated with Forensic mtDNA Typing. What is mtDNA Typing? Database and statistical issues General unacceptance. VALIDATION OF LABORATORY CONTAMINATION

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Mitochondrial DNA

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  1. Mitochondrial DNA • Observations on Frye/Daubert Issues Associated with Forensic mtDNA Typing. • What is mtDNA Typing? • Database and statistical issues • General unacceptance

  2. VALIDATION OF LABORATORY CONTAMINATION “Because of the sensitivity of this method. The issue of detection and quantitation of external contamination is important. We have observed low levels of amplified product in negative controls and reagent blanks when amplifying human mtDNA.” (Wilson et al., 1995a, p 667). “Previously sequenced amplicons were quantified by CE and two samples with different mtDNA control region sequences were chosen for this study. Amplified DNA was mixed together from the two samples in ratios of 4:1, 8:1, 12:1 and 16:1. Sequencing reactions were performed on the mixtures as previously described. Editing of the analyzed data from the sequence of the mixed samples was conducted by two individuals. Ambiguous bases (bases which could not be designated) and errors (bases which were called incorrectly) were noted. In the 4:1 mixture, both ambiguities and errors were observed. The 8:1 mixture yielded only two ambiguous calls from both editors at positions that differed in sequence between the mixed templates. No errors or ambiguities were noted at the 8:1 mixture. All base calls were completely consistent with the more abundant sequence at and above 8:1.” (Wilson et al., 1995a, p 667).

  3. LABORATORY CONTAMINATION, CONT. “In order to confirm that correct typing results can be achieved using this ratio, additional tests were conducted. Amplified DNA samples from a variety of donors were mixed in 10:1 ratios with other amplified DNAs. The dominant and minor samples were blindly reversed to the sequence editors. At the 10:1 ratio in a total of five such tests, all of the base calls from both editors were consistent with the known sequence of the more abundant sample with no errors or ambiguous calls.” (Wilson et al., 1995a, p 668).

  4. SEQUENCE FREQUENCY “Unless the discriminatory potential of a test can be objectively evaluated, an inclusion could mean anything. It is therefore incumbent on the forensic scientist to determine a means to evaluate and communicate the significance of an mtDNA inclusion or ‘match’.” Holland & Parsons, 1999. Forens. Sci. Intl.

  5. Native American Comparisons

  6. Asian Comparisons

  7. African & Pacific Group Comparisons

  8. Overall Search Results within Forensic Profiles Average Number of Differences = 6.681

  9. African Origin Database(s) within Forensic Profiles Average Number of Differences = 9.881

  10. Caucasian Origin Database(s) within Forensic Profiles Average Number of Differences = 4.111

  11. A 95% Upper Confidence Interval Using the Normal Approximation of the Binomial P  1.96 [PQ/N]1/2 ; where P = X/N, Q = 1-P, N = Database Size, and X = number of times a matching sequence is found in the database. Frequency of 0 differences = 9.2% Frequency of 1 difference = 27.2%

  12. CONFIDENCE LIMIT FROM ZERO PROPORTION P limit = 1 – a – 1/N, where N = Database Size N Maximum Frequency 50 0.058 1 in 17 100 0.030 1 in 34 200 0.015 1 in 67 500 0.006 1 in 167 1000 0.003 1 in 334 5000 0.0006 1 in 1670

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