An investigation of the rigor of interpretation rules for STRs derived from less than 100 pg of DNA

An investigation of the rigor of interpretation rules for STRs derived from less than 100 pg of DNA By increasing the PCR amplification regime to 34 cycles, we have demonstrated that it is possible routinely to analyse <100 pg DNA. The success rate was not improved (without impairing quality) by increasing cycle number further. Compared to amplification of 1 ng DNA at 28 cycles, it was shown that increased imbalance of heterozygotes occurred, along with an increase in the size (peak area) of stutters. The analysis of mixtures by peak area measurement becomes increasingly difficult as the sample size is reduced. Laboratory-based contamination cannot be completely avoided, even when analysis is carried out under stringent conditions of cleanliness. A set of guidelines that utilises duplication of results to interpret profiles originating from picogram levels of DNA is introduced. We demonstrate that the duplication guideline is robust by applying a statistical theory that models three key parameters — namely the incidence of allele drop-out, laboratory contamination and stutter. The advantage of the model is that the critical levels for each parameter can be calculated. This information may be used (for example) to determine levels of contamination that can be tolerated within the strategy employed. In addition we demonstrate that interpreting one banded loci, where allele dropout could have occurred, using LR =1/2 f a was conservative provided that the band was low in peak area. Furthermore, we demonstrate that an apparent mis-match between crime-stain and a suspect DNA profile does not necessarily result in an exclusion. The method used is complex, yet can be converted into an expert system. We envisage this to be the next step. http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png Forensic Science International Elsevier

An investigation of the rigor of interpretation rules for STRs derived from less than 100 pg of DNA

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Publisher
Elsevier
Copyright
Copyright © 2000 Elsevier Science Ireland Ltd
ISSN
0379-0738
DOI
10.1016/S0379-0738(00)00158-4
Publisher site
See Article on Publisher Site

Abstract

By increasing the PCR amplification regime to 34 cycles, we have demonstrated that it is possible routinely to analyse <100 pg DNA. The success rate was not improved (without impairing quality) by increasing cycle number further. Compared to amplification of 1 ng DNA at 28 cycles, it was shown that increased imbalance of heterozygotes occurred, along with an increase in the size (peak area) of stutters. The analysis of mixtures by peak area measurement becomes increasingly difficult as the sample size is reduced. Laboratory-based contamination cannot be completely avoided, even when analysis is carried out under stringent conditions of cleanliness. A set of guidelines that utilises duplication of results to interpret profiles originating from picogram levels of DNA is introduced. We demonstrate that the duplication guideline is robust by applying a statistical theory that models three key parameters — namely the incidence of allele drop-out, laboratory contamination and stutter. The advantage of the model is that the critical levels for each parameter can be calculated. This information may be used (for example) to determine levels of contamination that can be tolerated within the strategy employed. In addition we demonstrate that interpreting one banded loci, where allele dropout could have occurred, using LR =1/2 f a was conservative provided that the band was low in peak area. Furthermore, we demonstrate that an apparent mis-match between crime-stain and a suspect DNA profile does not necessarily result in an exclusion. The method used is complex, yet can be converted into an expert system. We envisage this to be the next step.

Journal

Forensic Science InternationalElsevier

Published: Jul 24, 2000

References

  • Analysis and interpretation of mixed forensic stains using DNA STR profiling
    Clayton, T.M.; Whitaker, J.P.; Sparkes, R.; Gill, P.
  • Development of guidelines to designate alleles using a STR multiplex system
    Gill, P.; Sparkes, R.; Kimpton, C.
  • Interpreting simple STR mixtures using allele peak area
    Gill, P.; Sparkes, R.; Pinchin, R.; Clayton, T.M.; Whitaker, J.P.; Buckleton, J.

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