A microhap panel for kinship analysis through massively parallel sequencing technology

Abstract

A microhap panel for kinship analysis through massively parallel sequencing technologyIt is widely recognized that microhaps are powerful markers for different forensic purposes, mainly due to their advantages of both short tandem repeats and single nucleotide polymorphisms, including multiple alleles, low mutation rate, and absence of stutter peaks. In the present study, a panel of 60 microhap loci was developed and utilized in forensic kinship analysis as a preliminary study. Genotyping of microhap was performed by massively parallel sequencing and haplotypes were directly achieved from sequence reads of 73 samples from Chinese Han population. We observed that 49 out of 60 loci have effective number of alleles greater than 3.0 and 10 out of 60 have values above 4.0, with an average value of 3.5598. The heterozygosity values were in a range from 0.5840 to 0.8546 with an average of 0.7268 and the cumulative power of exclusion value of the 60 loci is equal to 1-4.78 × 10 −18 . Moreover, we demonstrated the applicability of this method by different relationship inference problems, including identification of single parent-offspring, full-sibling, and second-degree relative. The results indicated that the assembled microhap panel provided more power for relationship inference, than commonly used short tandem repeats or single nucleotide polymorphism system.Abbreviations: Ae, effective number of alleles; LR, likelihood ratio; MH, microhap; MPS, massively parallel sequencing; PD, power of discriminative; PE, power of exclusion; PIC, polymorphism information content; ROI, region of interest; TPI, typical paternity index * These authors contributed equally to this work.Color online: See the article online to view Figs. 1-6 in color.

Evaluation of a Microhaplotype-Based Noninvasive Prenatal Test in Twin Gestations: Determination of Paternity, Zygosity, and Fetal Fraction

“…Sambamba tools were used to remove duplicate reads [26]. Microhaplotype alleles were assigned by an in-house Excel-based workbook as previously described [23].…”

“…A drop-in probability c of 0.01 and mutation rate r = 10 −8 were chosen, with silent allele s and theta correction θ set to 0. Allele frequency data for LR calculation are shown in our previous study [23], and a frequency of 0.002 ( 1 208×2+1 ) was applied for alleles not included in the data set.…”

“…To determine paternity, the relMix package was used to calculate the LRs that considered a few competing hypotheses as described above for each microhaplotype locus for each alleged father. Cumulative LR was attained by multiplying the individual LR values based on previous observations that the 60 loci are independent of each other [23]. As a result, the attained LR H1,H3 and LR H1,H2 values were all above 100 (in log 10 , ranging from 2.212 to 30.001) for 18 cases except case 2, with an assumed equal dropout probability d for both twins (ranging from 0.001 to 0.9), while the calculated values of LR H1,H3 and LR H2,H3 for case 2 were all below 1 (Table S1, Figure 2), indicating the inclusion of AFs as a biological father in relation to both twins in 18 cases but the exclusion of AF with regard to both twins in case 2.…”

“…However, little is known about the additional benefits of microhaplotypes in NIPT. In our previous study, a solution-based, high-redundancy DNA probe capture assay targeting 60 microhaplotypes was developed [22], demonstrating desirable performance in cfDNA sequencing and subsequent relationship interference in singleton pregnancies [23]. In this preliminary study, we evaluate the performance of microhaplotype-based NIPT to assign paternity, zygosity, and fetal fraction in twin gestations.…”

Considering the flanking region variants of nonbinary SNP and phenotype‐informative SNP to constitute 30 microhaplotype loci for increasing the discriminative ability of forensic applications

“…So, the microhaplotype loci established on the basis of nonbinary SNP and piSNP may be very useful forensic multifunctional markers. However, most other previously published microhaplotype panels were focused on the flanking region variations of the ancestry-informative SNP (aiSNP) and binary iiSNP [15][16][17][18][19][20][21][22][23][24][25][26][27][28][29][30]. So, the increased polymorphism and forensic applications of microhaplotype loci based on nonbinary SNP and piSNP should be further investigated.…”