A SNaPshot of next generation sequencing for forensic SNP analysis

Runa Daniel, Carla Santos, Christopher Phillips, Manuel Fondevila, Roland Van Oorschot, Angel Carracedo, Maria Lareu, Dennis MCNEVIN

Research output: Contribution to journalArticle

46 Citations (Scopus)

Abstract

Forensic phenotyping can provide useful intelligence regarding the biogeographical ancestry (BGA) and externally visible characteristics (EVCs) of the donor of an evidentiary sample. Currently, single nucleotide polymorphism (SNP) based inference of BGA and EVCs is performed most commonly using SNaPshot®, a single base extension (SBE) assay. However, a single SNaPshot multiplex PCR is limited to 30-40 SNPs. Next generation sequencing (NGS) offers the potential to genotype hundreds to thousands of SNPs from multiple samples in a single experimental run. The PCR multiplexes from five SNaPshot assays (SNPforID 52plex, SNPforID 34plex, Eurasiaplex, IrisPlex and an unpublished BGA assay) were applied to three different DNA template amounts (0.1, 0.2 and 0.3 ng) in three samples (9947A and 007 control DNAs and a male donor). The pooled PCR amplicons containing 136 unique SNPs were sequenced using Life Technologies' Ion Torrent™ PGM system. Approximately 72 Mb of sequence was generated from two 10 Mb Ion 314™ v1 chips. Accurate genotypes were readily obtained from all three template amounts. Of a total of 408 genotypes, 395 (97%) were fully concordant with SNaPshot across all three template amounts. Of those genotypes discordant with SNaPshot, six Ion Torrent sequences (1.5%) were fully concordant with Sanger sequencing across the three template amounts. Seven SNPs (1.7%) were either discordant between template amounts or discordant with Sanger sequencing. Sequence coverage observed in the negative control, and, allele coverage variation for heterozygous genotypes highlights the need to establish a threshold for background levels of sequence output and heterozygous balance. This preliminary study of the Ion Torrent PGM system has demonstrated considerable potential for use in forensic DNA analyses as a low to medium throughput NGS platform using established SNaPshot assays.
Original languageEnglish
Pages (from-to)50-60
Number of pages11
JournalForensic Science International: Genetics
Volume14
Issue number4
DOIs
Publication statusPublished - 1 Jan 2015

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Single Nucleotide Polymorphism
Genotype
Ions
Multiplex Polymerase Chain Reaction
DNA
Intelligence
Alleles
Technology
Polymerase Chain Reaction

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Daniel, R., Santos, C., Phillips, C., Fondevila, M., Van Oorschot, R., Carracedo, A., ... MCNEVIN, D. (2015). A SNaPshot of next generation sequencing for forensic SNP analysis. Forensic Science International: Genetics, 14(4), 50-60. https://doi.org/10.1016/j.fsigen.2014.08.013
Daniel, Runa ; Santos, Carla ; Phillips, Christopher ; Fondevila, Manuel ; Van Oorschot, Roland ; Carracedo, Angel ; Lareu, Maria ; MCNEVIN, Dennis. / A SNaPshot of next generation sequencing for forensic SNP analysis. In: Forensic Science International: Genetics. 2015 ; Vol. 14, No. 4. pp. 50-60.
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abstract = "Forensic phenotyping can provide useful intelligence regarding the biogeographical ancestry (BGA) and externally visible characteristics (EVCs) of the donor of an evidentiary sample. Currently, single nucleotide polymorphism (SNP) based inference of BGA and EVCs is performed most commonly using SNaPshot{\circledR}, a single base extension (SBE) assay. However, a single SNaPshot multiplex PCR is limited to 30-40 SNPs. Next generation sequencing (NGS) offers the potential to genotype hundreds to thousands of SNPs from multiple samples in a single experimental run. The PCR multiplexes from five SNaPshot assays (SNPforID 52plex, SNPforID 34plex, Eurasiaplex, IrisPlex and an unpublished BGA assay) were applied to three different DNA template amounts (0.1, 0.2 and 0.3 ng) in three samples (9947A and 007 control DNAs and a male donor). The pooled PCR amplicons containing 136 unique SNPs were sequenced using Life Technologies' Ion Torrent™ PGM system. Approximately 72 Mb of sequence was generated from two 10 Mb Ion 314™ v1 chips. Accurate genotypes were readily obtained from all three template amounts. Of a total of 408 genotypes, 395 (97{\%}) were fully concordant with SNaPshot across all three template amounts. Of those genotypes discordant with SNaPshot, six Ion Torrent sequences (1.5{\%}) were fully concordant with Sanger sequencing across the three template amounts. Seven SNPs (1.7{\%}) were either discordant between template amounts or discordant with Sanger sequencing. Sequence coverage observed in the negative control, and, allele coverage variation for heterozygous genotypes highlights the need to establish a threshold for background levels of sequence output and heterozygous balance. This preliminary study of the Ion Torrent PGM system has demonstrated considerable potential for use in forensic DNA analyses as a low to medium throughput NGS platform using established SNaPshot assays.",
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Daniel, R, Santos, C, Phillips, C, Fondevila, M, Van Oorschot, R, Carracedo, A, Lareu, M & MCNEVIN, D 2015, 'A SNaPshot of next generation sequencing for forensic SNP analysis', Forensic Science International: Genetics, vol. 14, no. 4, pp. 50-60. https://doi.org/10.1016/j.fsigen.2014.08.013

A SNaPshot of next generation sequencing for forensic SNP analysis. / Daniel, Runa; Santos, Carla; Phillips, Christopher; Fondevila, Manuel; Van Oorschot, Roland; Carracedo, Angel; Lareu, Maria; MCNEVIN, Dennis.

In: Forensic Science International: Genetics, Vol. 14, No. 4, 01.01.2015, p. 50-60.

Research output: Contribution to journalArticle

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T1 - A SNaPshot of next generation sequencing for forensic SNP analysis

AU - Daniel, Runa

AU - Santos, Carla

AU - Phillips, Christopher

AU - Fondevila, Manuel

AU - Van Oorschot, Roland

AU - Carracedo, Angel

AU - Lareu, Maria

AU - MCNEVIN, Dennis

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Daniel R, Santos C, Phillips C, Fondevila M, Van Oorschot R, Carracedo A et al. A SNaPshot of next generation sequencing for forensic SNP analysis. Forensic Science International: Genetics. 2015 Jan 1;14(4):50-60. https://doi.org/10.1016/j.fsigen.2014.08.013