Evaluating 130 microhaplotypes across a global set of 83 populations.

Today the primary DNA markers used in forensics are short tandem repeat (STR) polymorphisms (STRPs), initially selected because they are highly polymorphic. However, the increasingly common need to deal with samples with a mixture of DNA from two or more individuals sometimes is complicated by the inherent stutter involved with PCR amplification, especially in strongly unbalanced mixtures when the minor component coincides with the stutter range of the major component. Also, the STRPs in use provide little evidence of ancestry of a single source sample beyond broad "continental" resolution. Methodologies for analyzing DNA have become much more powerful in recent years. Massively parallel sequencing (MPS) is a new method being considered for routine use in forensics. Primarily to aid in mixture deconvolution and avoid the issue of stutter, we have begun to investigate a new type of forensic marker, microhaplotype loci, that will provide useful information on mixtures of DNA and on ancestry when typed using massively parallel sequencing (MPS). We have identified 130 loci and estimated their haplotype (allele) frequencies in 83 different population samples. Many of these loci are shown to be highly informative for individual identification and for mixture identification and deconvolution.

A 50-SNP assay for biogeographic ancestry and phenotype prediction in the U.S. population.

When an STR DNA profile obtained from crime scene evidence does not match identified suspects or profiles from available databases, further DNA analyses targeted at inferring the possible ancestral origin and phenotypic characteristics of the perpetrator could yield valuable information. Single Nucleotide Polymorphisms (SNPs), the most common form of genetic polymorphisms, have alleles associated with specific populations and/or correlated to physical characteristics. We have used single base primer extension (SBE) technology to develop a 50 SNP assay (composed of three multiplexes) designed to predict ancestry among the primary U.S. populations (African American, East Asian, European American, and Hispanic American/Native American), as well as pigmentation phenotype (eye, hair, and skin color) among European American. We have optimized this assay to a sensitivity level comparable to current forensic DNA analyses, and shown robust performance on forensic-type samples. In addition, we developed a prediction model for ancestry in the U.S. population, based on the random match probability and likelihood ratio formulas already used in forensic laboratories. Lastly, we evaluated the biogeographic ancestry prediction model using a test set, and we evaluated an existing model for eye color with our U.S. sample set. Using these models with recommended thresholds, the 50 SNP assay provided accurate ancestry information in 98.6% of the test set samples, and provided accurate eye color information in 61% of the European samples tested (25% were inconclusive and 14% were incorrect). This method, which uses equipment already available in forensic DNA laboratories, is recommended for use in U.S. forensic casework to provide additional information about the donor of a DNA sample when the STR profile has not been linked to an individual.