STR Interpretation 2

STR Interpretation 2 - Routine challenging features of...

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1. Forensic samples of minimal quantity. 2. Mixed samples. Environmental Insults -Conditions that evidentiary samples are often exposed to can degrade DNA. -Degradation affects loci differently based upon amplicon size. Degraded template (DNA) RFU Locus 1 Locus 2 Locus 3 Threshold Increasing size of PCR product Peak(s) below threshold due to DNA degradation Result of degradation: Effective reduction in the number of template strands for the larger loci decreases amplification product. = Potential loss of larger loci. When Amplicon Size is Critical The effective template concentration varies from locus to locus based upon amplicon size. With degraded samples, all loci do not behave the same. With non-degraded samples (HMW DNA), all loci behave ~ the same. If one amplifies, they all amplify. However, Example of Degraded Sample
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When are minor peaks true alleles? The location of minor peak(s) can be a factor in the interpretation of the results. Is the minor peak in the stutter position a true allele or just stutter? 20 % The empirically derived stutter percentage data (user lab or kit manufacturer) is used in the interpretation of this scenario . Example: If the N-4 peak is 20% of the major allelic peak and the stutter threshold for this allele is 10%, this indicates that the N-4 peak is a true allele (esp. if mixture is clearly shown at other loci). Minor peaks continued Is the shoulder peak a minor component to a mixture or machine noise/artifact? Interpretation is based on how many RFU the peak is above the shoulder. If peak is only ~ 10 RFUs above shoulder = not called. Minor peak? If true peak is 75+ RFU (factor the effect of the shoulder = called an allele. Stochastic Fluctuation •Stochastic = chance. •Result of PCR founder effect and chance allele Sampling. If you amplify small amounts of DNA, can see stochastic effects. Results in Stochastic Zone
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This note was uploaded on 09/21/2009 for the course MCB 3416 taught by Professor Ladd during the Fall '08 term at UConn.

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STR Interpretation 2 - Routine challenging features of...

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