Developmental Validation of a DIP-InDel/DIP-STR System for the Detection of Unbalanced DNA Mixtures

Extracting "masked" minor donor information from an unbalanced DNA mixture stain is important for the identification of the person of interest. In recent years, a series of compound markers genotyped based on allele-specific amplifications have been proposed to detect minor contributors of mixed stains. In this study, we selected out 18 DIP-InDel markers from previous literatures and established a multiplex system encompassing 18 DIP-InDel markers and 6 DIP-STR markers in two separate reactions. The allele frequencies were estimated based on 200 samples, and 23 of the 24 DIP-linked length polymorphic markers had a relatively high probability of informative markers (I value >0.267), which indicated their potential usefulness in the Chinese Han population. Moreover, the multiplex was highly sensitive (requiring >0.025 ng) and specific to human DNA. This system is capable of detecting the minor contributor comprising 1% in unbalanced mixtures of two individuals. The capacity of profiling cell-free DNA was also analyzed in few cases. At least one paternal allele could be detected in maternal plasma samples with gestation periods ranging from 27 to 40 weeks. To conclude, the DIP-linked length polymorphic markers may serve as a convenient method for detecting the presence of a minor donor in unbalanced mixtures and show promise for noninvasive prenatal diagnosis.