Individual identification and kinship testing from hair shaft nuclear DNA: leveraging short amplicon strategy and bioinformatics models.

Hair samples are commonly encountered at crime scenes. However, most of them are telogen hairs, from which only partial DNA profiles can be yielded. These profiles are generally insufficient for forensic human identification and kinship testing. In this study, we sequenced the hair roots and hair shafts from ten unrelated individuals using a short-amplicon panel, and evaluated the performance of hair shafts of different lengths in terms of genotype calling and genotype accuracy. We also employed a pseudohaploid genotype-based approach for kinship coefficient estimation and proposed an error model for likelihood ratio (LR) calculation. The results showed that many SNPs could be successfully recovered from the nuclear DNA of hair shafts. However, approximately 30% of the genotypes called were inconsistent with those of the hair roots, 17.32%, 71.88%, and 10.79% of which were drop-in, dropout, and switch errors, respectively. Five hair shafts were found to be contaminated by either the experimental operator or by other samples. A significant decrease in kinship coefficients was observed for related individuals with short hairs, while there was no significant difference for unrelated pairs. The overall accuracy was 55.15% for identical samples, parent-child relationships, and grandparent-grandchild relationships, which is 4.55 times that using KING. In terms of the LR model, the average effectiveness was 0.63, 0.41, and 0.32 for individual identification, paternity testing, and grandparent-grandchild testing, respectively when a 2 cm hair shaft was used; these values increased to 0.95, 0.93, and 0.74 when a 15 cm hair shaft was used. In addition, no unrelated pairs were incorrectly judged as related for individual identification and paternity testing. In conclusion, the short amplicon strategy and our proposed bioinformatics methods make it possible to perform human identification and kinship testing using nuclear DNA from hair shafts. However, contamination poses a substantial challenge when working with such low-quality DNA.