法生物学的試料由来のＤＮＡに対する定量法の検証

Abstract

A quantitative PCR procedure using a newly designed primer set for amplification of human specific DNA sequence D17Z1 for quantification of human DNA has been developed. The procedure was compared with the existing quantification method, UV absorption method and slot blot hybridization method. The lower limit of quantification, the reproducibility of results, quantification of degraded DNA, quantification specificity and application for forensic casework samples were examined using each technique. The quantitative PCR method consumed the least amount of DNA and could quantify as little as 0.0015 ng of DNA, and its reproducibility was comparable with the UV absorption method. The STR typing of degraded DNA samples quantified by slot blot hybridization method and quantitative PCR method were properly detected and typed. Since the bacterial DNA samples were not detected and quantified by slot blot hybridization method and quantitative PCR method, they are useful for human-specific DNA quantification of forensic evidence samples containing bacterial DNA. In the experimental application of the quantitative PCR method for various forensic casework samples, almost every STR loci were properly detected and it demonstrated a beneficial effect for actual forensic identification analysis. Furthermore, the quantitative PCR method was also useful in determining the proper amount of template DNA extracted from the sample suspected to include amplification inhibitor. In the results of the comparisons, the human specific DNA quantification using the technique of quantitative PCR is more practical for forensic DNA analysis than the other methods.