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 demonstrat ed that it is possible routinely to analyse <100 pg DNA, The success rate w as not improved (without impairing quality) by increasing cycle number furt her. Compared to amplification of 1 ng DNA at 28 cycles, it was shown that increased imbalance of heterozygotes occurred, along with an increase in th e size (peak area) of stutters. The analysis of mixtures by peak area measu rement becomes increasingly difficult as the sample size is reduced. Labora tory-based contamination cannot be completely avoided, even when analysis i s carried out under stringent conditions of cleanliness. A set of guideline s that utilises duplication of results to interpret profiles originating fr om picogram levels of DNA is introduced. We demonstrate that the duplicatio n guideline is robust by applying a statistical theory that models three ke y parameters - namely the incidence of allele drop-out, laboratory contamin ation and stutter. The advantage of the model is that the critical levels f or each parameter can be calculated. This information may be used (for exam ple) 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 LX = 1/2f(a) was cons ervative provided that the band was low in peak area. Furthermore, we demon strate that an apparent mis-match between crime-stain and a suspect DNA pro file does not necessarily result in an exclusion. The method used is comple x, yet can be converted into an expert system. We envisage this to be the n ext step. (C) 2000 Elsevier Science Ireland Ltd. All rights reserved.