Development and Validation of a Forensic Multiplex System With 38 X-InDel Loci.

Ling Chen,Weian Du,Weibin Wu,Changhui Liu,Zhenya Tang,Xiyong Pan,Chao Liu,Xiaolong Han,Cheng Xiao,Yuan Wang

doi:10.3389/fgene.2021.670482

Ling Chen, Weian Du + Show 8 more

Open Access

https://doi.org/10.3389/fgene.2021.670482

Copy DOI

Abstract

In the present study, a novel multiplex system, AGCU X-InDel 38 kit, was designed to amplify 38 X-InDel markers and amelogenin in a single Polymerase Chain Reaction (PCR). To demonstrate the suitability and efficiency for forensic applications, a series of validation experiments were conducted, including sensitivity, species specificity, reproducibility, stability, case samples, balance of peak height, size precision, as well as allele frequency and forensic parameter analysis. The results showed that AGCU X-InDel 38 kit was capable to get full profiles even with 62.5 pg of template DNA, and full profiles can be obtained when hematin concentration ≤25 μmol/L, or hemoglobin concentration ≤50 μmol/L, showing good tolerance to six common inhibitors. Moreover, the analyzed case samples indicated that AGCU X-InDel 38 kit had better performance for degraded and trace DNA samples. The 200 unrelated males from Guangdong Han population showed that the combined PDMale and PDFemale were both more than 0.999999999, and the combined MECKrüger, MECKishida, and MECDesmarais Duo were 0.999369481, 0.999999917, and 0.999941556, respectively. Robust discrimination capability of this novel multiplex system could be demonstrated through the high values of forensic parameters. In conclusion, AGCU X-InDel 38 kit is sensitive, precise, reproducible, and highly informative and could be used as a complementary tool for complex and challenging kinship cases.

Highlights

Short tandem repeat (STR) loci represent the mainstream markers for forensic investigations
According to sequences from the dbSNP database, X-InDel markers were selected to meet the following criteria: (1) the difference of allelic X-InDel fragment in length, the inserted or deleted base number in other words, ranged from 1 to 25 bp; (2) all loci were located in introns; (3) the minor allele frequency (MAF) varied from 0.4 to 0.5 in East Asians; and (4) all loci comply with the Hardy–Weinberg equilibrium in the Chinese population
WD and XH helped perform the analysis with constructive discussions

Summary

Introduction

Short tandem repeat (STR) loci represent the mainstream markers for forensic investigations. Autosomal STRs may not be effective in some deficiency paternity cases. Taking advantages of X-linked inheritance into consideration, X chromosome markers are of great value in some specific and deficient kinship cases especially when autosomal chromosome markers. Different from STRs, InDel markers have a short amplicon size and low mutation rates, and simple analytical procedures are required (LaRue et al, 2012; Chen et al, 2019). There is a growing tendency in studying X chromosomal InDel markers, especially in the field of evolutionary anthropology, assessing admixture of population and kinship investigations with deficient relationship (Ribeiro-Rodrigues et al, 2009; Resque et al, 2010; Ibarra et al, 2014)

Methods

Results

Conclusion