Abstract
Like DIP-STR markers (deletion/insertion polymorphism-short tandem repeat combinations), SNP-STR markers (single nucleotide polymorphism-STR combinations) are also valuable in forensic DNA mixture analysis. In this study, eight SNP-STRs were selected, and a stable and sensitive multiplex polymerase chain reaction (PCR) assay was developed for amplifying these SNP-STRs and the Amelogenin gender marker according to the principle of amplification refractory mutation system (ARMS). This novel multiplex set allows detection of the minor DNA contributor in a DNA mixture of any gender and cellular origin with high resolution (beyond a DNA ratio of 1:20). In addition, SNP-STR haplotype frequencies were estimated based on a survey of 350 unrelated individuals from Chinese Han population, and the combined power of discrimination (PD) and power of exclusion (PE) of the eight SNP-STRs were calculated as 0.99999999965 and 0.9996, which were obviously higher than that of the eight STR loci: 0.9999999954 and 0.9989 respectively. The results indicated that the SNP-STR compound markers have higher application value in forensic identification compared to standard autosomal STRs, especially in the analysis of imbalanced DNA mixtures.
Highlights
Mixed stains derived from different contributors are common biological evidence samples in forensic practice, and these complex biological samples generate mixed genotypes, presenting challenges in interpreting the results, especially for those imbalanced genomic mixtures [1, 2]
Six of the eight single nucleotide polymorphisms amplified with STRs (SNP-STR) locate on different chromosomes, and the other two markers, rs1276598-D6S474 and rs2325399-D6S1043, locate on the same chromosomal arm (6q), and their physical distance and genetic distance are about 20.8 Mb and 17.73 cM (Marshfield) respectively, the pairwise linkage disequilibrium analysis showed that the two markers were genetically independent (p = 0.1675) in the studied Chinese Han population
For the SNPs, the minor allele frequencies of all selected loci in Chinese Southern Han population is higher than 0.2 except for the rs16887642 (0.1857) according to 1000 genomes databases. This multiplex set was designed as a 5-dye assay, two SNP allele-specific primers for each SNP-STR marker were labeled at the 50-end respectively with 6-FAM and HEX or TAMRA and ROX fluorescent dye for the detection by ABI 3130 Genetic Analyzer (Thermo Fisher Scientific, MA, USA)
Summary
Mixed stains derived from different contributors are common biological evidence samples in forensic practice, and these complex biological samples generate mixed genotypes, presenting challenges in interpreting the results, especially for those imbalanced genomic mixtures [1, 2]. As the common forensic DNA analysis method, one of the limitations of the capillary electrophoresis (CE)-based polymerase chain reaction (PCR)-STR typing technique is that it does not work successfully if the proportion of the DNA quantities of the two contributors is more extreme than 1:10 [3]. Y-chromosome STRs can be used to detect the male component in these mixed samples when the DNA of the male contributor is present in a small amount [4]. A variety of strategies have been developed to separate different cell populations prior.
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