Comparison of CE- and MPS-based analyses of forensic markers in a single cell after whole genome amplification

Abstract

Whole genome amplification (WGA) allows for multiple genetic analyses with low template DNA, such as DNA derived from a single cell. WGA could increase the amount of input DNA from the pg to the μg level. However, there are no studies comparing the performance of forensic markers with DNA from a single cell after WGA evaluated on both capillary electrophoresis (CE) and massively parallel sequencing (MPS) platforms. In this study, cell lines consisting of female cultured B-lymphoblastoid cells and karyocytes from male venous blood were segregated into one, two, three and five cells. Including the references with the bulk cells, all samples were generated by WGA with the multiple displacement amplification (MDA) strategy in triplicate and genotyped on CE and MPS platforms. Allele balance, stutter ratio, accuracy, repeatability and concordance of short tandem repeat (STR) markers were used to evaluate the genotyping performance on both platforms. Additionally, the sequence coverage ratio (SCR) and SNP genotypes were evaluated for sequence information generated from the MPS. Heterozygous loci showed high allele balance, with an overall average allele balance ratio larger than 0.79 on the CE and 0.75 on the MPS platforms for the venous blood cell samples; the cultured B-lymphoblastoid cell samples had ratios of 0.62 and 0.70, respectively. The stutter ratio of every source and cell number from both cell line samples were very close, ranging from 5.3%-7.2% for autosomal STRs and approximately 10 % of Y chromosomal STRs on the CE platform. The average stutter, allele, and sequence-based and length-based noise ratios were 6.6 %, 88 %, 4.7 % and 0.7 %, respectively, in the single male cell sample. SNPs also showed high consistency and intralocus balance. Our study indicated that WGA with MDA strategy works relatively well of STR and SNP genotyping with low copy number samples on CE and MPS, even with one-cell sample.

Keywords: Capillary electrophoresis; Forensic loci; Low copy number; Massively parallel sequencing; Whole genome amplification.