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High sensitivity multiplex short tandem repeat loci analyses with massively parallel sequencing.
Forensic Sci Int Genet. 2015 May; 16:38-47.FS

Abstract

STR typing in forensic genetics has been performed traditionally using capillary electrophoresis (CE). However, CE-based method has some limitations: a small number of STR loci can be used; stutter products, dye artifacts and low level alleles. Massively parallel sequencing (MPS) has been considered a viable technology in recent years allowing high-throughput coverage at a relatively affordable price. Some of the CE-based limitations may be overcome with the application of MPS. In this study, a prototype multiplex STR System (Promega) was amplified and prepared using the TruSeq DNA LT Sample Preparation Kit (Illumina) in 24 samples. Results showed that the MinElute PCR Purification Kit (Qiagen) was a better size selection method compared with recommended diluted bead mixtures. The library input sensitivity study showed that a wide range of amplicon product (6-200ng) could be used for library preparation without apparent differences in the STR profile. PCR sensitivity study indicated that 62pg may be minimum input amount for generating complete profiles. Reliability study results on 24 different individuals showed that high depth of coverage (DoC) and balanced heterozygote allele coverage ratios (ACRs) could be obtained with 250pg of input DNA, and 62pg could generate complete or nearly complete profiles. These studies indicate that this STR multiplex system and the Illumina MiSeq can generate reliable STR profiles at a sensitivity level that competes with current widely used CE-based method.

Authors+Show Affiliations

Institute of Applied Genetics, Department of Molecular and Medical Genetics, University of North Texas Health Science Center, 3500 Camp Bowie Blvd., Fort Worth, TX 76107, USA.Institute of Applied Genetics, Department of Molecular and Medical Genetics, University of North Texas Health Science Center, 3500 Camp Bowie Blvd., Fort Worth, TX 76107, USA.Institute of Applied Genetics, Department of Molecular and Medical Genetics, University of North Texas Health Science Center, 3500 Camp Bowie Blvd., Fort Worth, TX 76107, USA.Institute of Applied Genetics, Department of Molecular and Medical Genetics, University of North Texas Health Science Center, 3500 Camp Bowie Blvd., Fort Worth, TX 76107, USA.Institute of Applied Genetics, Department of Molecular and Medical Genetics, University of North Texas Health Science Center, 3500 Camp Bowie Blvd., Fort Worth, TX 76107, USA.Institute of Applied Genetics, Department of Molecular and Medical Genetics, University of North Texas Health Science Center, 3500 Camp Bowie Blvd., Fort Worth, TX 76107, USA; Department of Forensic Medicine, Hjelt Institute, University of Helsinki, P.O. Box 40, 00014 Helsinki, Finland.Promega Corporation, 2800 Woods Hollow Road, Madison, WI 53711, USA.Promega Corporation, 2800 Woods Hollow Road, Madison, WI 53711, USA.Institute of Applied Genetics, Department of Molecular and Medical Genetics, University of North Texas Health Science Center, 3500 Camp Bowie Blvd., Fort Worth, TX 76107, USA; Center of Excellence in Genomic Medicine Research (CEGMR), King Abdulaziz University, Jeddah, Saudi Arabia. Electronic address: Bruce.Budowle@unthsc.edu.

Pub Type(s)

Comparative Study
Journal Article
Research Support, U.S. Gov't, Non-P.H.S.

Language

eng

PubMed ID

25528025

Citation

Zeng, Xiangpei, et al. "High Sensitivity Multiplex Short Tandem Repeat Loci Analyses With Massively Parallel Sequencing." Forensic Science International. Genetics, vol. 16, 2015, pp. 38-47.
Zeng X, King JL, Stoljarova M, et al. High sensitivity multiplex short tandem repeat loci analyses with massively parallel sequencing. Forensic Sci Int Genet. 2015;16:38-47.
Zeng, X., King, J. L., Stoljarova, M., Warshauer, D. H., LaRue, B. L., Sajantila, A., Patel, J., Storts, D. R., & Budowle, B. (2015). High sensitivity multiplex short tandem repeat loci analyses with massively parallel sequencing. Forensic Science International. Genetics, 16, 38-47. https://doi.org/10.1016/j.fsigen.2014.11.022
Zeng X, et al. High Sensitivity Multiplex Short Tandem Repeat Loci Analyses With Massively Parallel Sequencing. Forensic Sci Int Genet. 2015;16:38-47. PubMed PMID: 25528025.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - High sensitivity multiplex short tandem repeat loci analyses with massively parallel sequencing. AU - Zeng,Xiangpei, AU - King,Jonathan L, AU - Stoljarova,Monika, AU - Warshauer,David H, AU - LaRue,Bobby L, AU - Sajantila,Antti, AU - Patel,Jaynish, AU - Storts,Douglas R, AU - Budowle,Bruce, Y1 - 2014/12/03/ PY - 2014/08/22/received PY - 2014/10/20/revised PY - 2014/11/26/accepted PY - 2014/12/22/entrez PY - 2014/12/22/pubmed PY - 2016/2/26/medline KW - Illumina MiSeq KW - Massively parallel sequencing (MPS) KW - Short tandem repeat (STR) SP - 38 EP - 47 JF - Forensic science international. Genetics JO - Forensic Sci Int Genet VL - 16 N2 - STR typing in forensic genetics has been performed traditionally using capillary electrophoresis (CE). However, CE-based method has some limitations: a small number of STR loci can be used; stutter products, dye artifacts and low level alleles. Massively parallel sequencing (MPS) has been considered a viable technology in recent years allowing high-throughput coverage at a relatively affordable price. Some of the CE-based limitations may be overcome with the application of MPS. In this study, a prototype multiplex STR System (Promega) was amplified and prepared using the TruSeq DNA LT Sample Preparation Kit (Illumina) in 24 samples. Results showed that the MinElute PCR Purification Kit (Qiagen) was a better size selection method compared with recommended diluted bead mixtures. The library input sensitivity study showed that a wide range of amplicon product (6-200ng) could be used for library preparation without apparent differences in the STR profile. PCR sensitivity study indicated that 62pg may be minimum input amount for generating complete profiles. Reliability study results on 24 different individuals showed that high depth of coverage (DoC) and balanced heterozygote allele coverage ratios (ACRs) could be obtained with 250pg of input DNA, and 62pg could generate complete or nearly complete profiles. These studies indicate that this STR multiplex system and the Illumina MiSeq can generate reliable STR profiles at a sensitivity level that competes with current widely used CE-based method. SN - 1878-0326 UR - https://www.unboundmedicine.com/medline/citation/25528025/High_sensitivity_multiplex_short_tandem_repeat_loci_analyses_with_massively_parallel_sequencing_ DB - PRIME DP - Unbound Medicine ER -