TY - JOUR
T1 - Sequence-based analysis of stutter at STR loci
T2 - Characterization and utility
AU - Aponte, Rachel A.
AU - Gettings, Katherine B.
AU - Duewer, David L.
AU - Coble, Michael D.
AU - Vallone, Peter M.
N1 - Funding Information:
This work was funded in part by the Federal Bureau of Investigation (FBI) interagency agreement DJF-13-0100-PR-0000080: “DNA as a Biometric”.
Publisher Copyright:
© 2015
Copyright:
Copyright 2017 Elsevier B.V., All rights reserved.
PY - 2015/12/1
Y1 - 2015/12/1
N2 - The development of next generation sequencing (NGS) technologies creates the potential for changing the method by which the forensic science community genotypes short tandem repeat (STR) loci. While the capabilities of NGS are promising, moving from current capillary electrophoresis (CE) methods would require new guidelines to be established and a new understanding of artifacts that may arise with the use of NGS. Stutter has been well characterized for CE technologies; however, NGS workflows may use different polymerases and amplification approaches, which could alter the appearance of this artifact. Stutter is most commonly seen in the n-4 position in CE data, but may be observed more rarely in the n+4 and n-8 positions. NGS data frequently contains detectable sequences consistent with stutter at the n+4 and n-8 positions, and may even contain stutter at the n-12 position for some loci. It is possible that these alternate types of stutter events occur at similar levels in CE workflows and go undetected due to the analytical threshold employed or because the artifacts do not exceed the background noise. Comparing stutter events in NGS data to what has been observed by CE will improve our understanding of the effects of library preparation and sequencing. Characterizing stutter events by sequence will contribute to the development of guidelines and facilitate implementation of NGS technology. Further, determining stutter ratios for each isoallele would allow for individual sequence thresholds to be set, which could then be used to improve mixture interpretation models.
AB - The development of next generation sequencing (NGS) technologies creates the potential for changing the method by which the forensic science community genotypes short tandem repeat (STR) loci. While the capabilities of NGS are promising, moving from current capillary electrophoresis (CE) methods would require new guidelines to be established and a new understanding of artifacts that may arise with the use of NGS. Stutter has been well characterized for CE technologies; however, NGS workflows may use different polymerases and amplification approaches, which could alter the appearance of this artifact. Stutter is most commonly seen in the n-4 position in CE data, but may be observed more rarely in the n+4 and n-8 positions. NGS data frequently contains detectable sequences consistent with stutter at the n+4 and n-8 positions, and may even contain stutter at the n-12 position for some loci. It is possible that these alternate types of stutter events occur at similar levels in CE workflows and go undetected due to the analytical threshold employed or because the artifacts do not exceed the background noise. Comparing stutter events in NGS data to what has been observed by CE will improve our understanding of the effects of library preparation and sequencing. Characterizing stutter events by sequence will contribute to the development of guidelines and facilitate implementation of NGS technology. Further, determining stutter ratios for each isoallele would allow for individual sequence thresholds to be set, which could then be used to improve mixture interpretation models.
KW - Next generation sequencing (NGS)
KW - Short tandem repeat (STR)
KW - Stutter
UR - http://www.scopus.com/inward/record.url?scp=84983503846&partnerID=8YFLogxK
U2 - 10.1016/j.fsigss.2015.09.181
DO - 10.1016/j.fsigss.2015.09.181
M3 - Article
AN - SCOPUS:84983503846
SN - 1875-1768
VL - 5
SP - e456-e458
JO - Forensic Science International: Genetics Supplement Series
JF - Forensic Science International: Genetics Supplement Series
ER -