Abstract
Tri-allelic SNP has been regarded as a potential marker in forensic utility. Compared with the conventional binary SNP marker, it has showed obvious advantage in the analysis of mixed and degraded DNA samples. Previously, we constructed a multiplex system with 20 tri-allelic SNP loci by the pyrosequencing (PSQ) method and SNaPshot technique. In the present work, validation studies were carried out to determine the efficacy and reliability of the multiplex assay for forensic identification. Species specificity, sensitivity, artificially degraded samples and mixture study were executed in our research, according to the FBI/National Standards and Scientific Working Group on DNA Analysis Methods (SWGDAM) guideline. The results demonstrated that the multiplex system was sensitive to 1ng of input DNA. The complete SNP profiles could be consistently detected from the degraded DNA. No profiles were obtained in the forensically relevant animals. Additionally, using the multiplex SNP system, the presence of a mixture of two DNA samples in a ratio up to 3:7 could be recognized reliably. In conclusion, the 20-tri-allelic SNP multiplex system was suitable for paternity testing and human identification in forensic science.
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