title: Developmental validation of a custom panel including 273 SNPs for forensic application using Ion Torrent PGM.
abstract: Utilizing massively parallel sequencing (MPS) technology for SNP testing in forensic genetics is becoming attractive because of the shortcomings of STR markers, such as their high mutation rates and disadvantages associated with the current PCR-CE method as well as its limitations regarding multiplex capabilities. MPS offers the potential to genotype hundreds to thousands of SNPs from multiple samples in a single experimental run. In this study, we designed a customized SNP panel that includes 273 forensically relevant identity SNPs chosen from SNPforID, IISNP, and the HapMap database as well as previously related studies and evaluated the levels of genotyping precision, sequence coverage, sensitivity and SNP performance using the Ion Torrent PGM. In a concordant study of the custom MPS-SNP panel, only four MPS callings were missing due to coverage reads that were too low (<20), whereas the others were fully concordant with Sanger's sequencing results across the two control samples, that is, 9947A and 9948. The analyses indicated a balanced coverage among the included loci, with the exception of the 16 SNPs that were used to detect an inconsistent allele balance and/or lower coverage reads among 50 tested individuals from the Chinese HAN population and the above controls. With the exception of the 16 poorly performing SNPs, the sequence coverage obtained was extensive for the bulk of the SNPs, and only three Y-SNPs (rs16980601, rs11096432, rs3900) showed a mean coverage below 1000. Analyses of the dilution series of control DNA 9948 yielded reproducible results down to 1ng of DNA input. In addition, we provide an analysis tool for automated data quality control and genotyping checks, and we conclude that the SNP targets are polymorphic and independent in the Chinese HAN population. In summary, the evaluation of the sensitivity, accuracy and genotyping performance provides strong support for the application of MPS technology in forensic SNP analysis, and the assay offers a straightforward sample-to-genotype workflow that could be beneficial in forensic casework with respect to both individual identification and complex kinship issues.
authors.affiliation: Shanghai Key Laboratory of Forensic Medicine, Shanghai Forensic Service Platform, Institute of Forensic Sciences, Ministry of Justice, Shanghai 200063, PR China.
authors.forename: Suhua
authors.initials: S
authors.lastname: Zhang
authors.affiliation: Shanghai Key Laboratory of Forensic Medicine, Shanghai Forensic Service Platform, Institute of Forensic Sciences, Ministry of Justice, Shanghai 200063, PR China.
authors.forename: Yingnan
authors.initials: Y
authors.lastname: Bian
authors.affiliation: State Key Laboratory of Genetic Engineering, Institute of Genetics, School of Life Sciences, Fudan University, Shanghai 200433, PR China.
authors.forename: Anqi
authors.initials: A
authors.lastname: Chen
authors.affiliation: Department of Forensic Medicine, Medical College of Soochow University, Suzhou 215123, PR China.
authors.forename: Hancheng
authors.initials: H
authors.lastname: Zheng
authors.affiliation: Department of Forensic Medicine, Medical College of Soochow University, Suzhou 215123, PR China.
authors.forename: Yuzhen
authors.initials: Y
authors.lastname: Gao
authors.affiliation: Department of Forensic Genetics, West China School of Preclinical and Forensic Medicine, Sichuan University, Chengdu 610041, PR China.
authors.forename: Yiping
authors.initials: Y
authors.lastname: Hou
authors.affiliation: Shanghai Key Laboratory of Forensic Medicine, Shanghai Forensic Service Platform, Institute of Forensic Sciences, Ministry of Justice, Shanghai 200063, PR China. Electronic address: lichengtaohla@163.com.
authors.forename: Chengtao
authors.initials: C
authors.lastname: Li
mesh_terms: D044466:Asian People / Q000235:genetics; D002681:China; D005006:Ethnicity / Q000235:genetics; D005260:Female; D005787:Gene Frequency; D005838:Genotype; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods*; D006801:Humans; D008297:Male; D060885:Multiplex Polymerase Chain Reaction; D020641:Polymorphism, Single Nucleotide; D012984:Software
keywords: Forensic Genetics; Massively Parallel Sequencing (MPS); Personal Genome Machine (PGM); SNP