title: Massively parallel multiplex DNA sequencing for specimen identification using an Illumina MiSeq platform.
abstract: Genetic information is a valuable component of biosystematics, especially specimen identification through the use of species-specific DNA barcodes. Although many genomics applications have shifted to High-Throughput Sequencing (HTS) or Next-Generation Sequencing (NGS) technologies, sample identification (e.g., via DNA barcoding) is still most often done with Sanger sequencing. Here, we present a scalable double dual-indexing approach using an Illumina Miseq platform to sequence DNA barcode markers. We achieved 97.3% success by using half of an Illumina Miseq flowcell to obtain 658 base pairs of the cytochrome c oxidase I DNA barcode in 1,010 specimens from eleven orders of arthropods. Our approach recovers a greater proportion of DNA barcode sequences from individuals than does conventional Sanger sequencing, while at the same time reducing both per specimen costs and labor time by nearly 80%. In addition, the use of HTS allows the recovery of multiple sequences per specimen, for deeper analysis of genetic variation in target gene regions.
authors.affiliation: Department of Integrative Biology and Biodiversity Institute of Ontario, University of Guelph, 50 Stone Road East, Guelph, ON, Canada N1G 2W1.
authors.forename: Shadi
authors.initials: S
authors.lastname: Shokralla
authors.affiliation: Department of Biology, McMaster University, 1280 Main Street West, Hamilton, ON, Canada L8S 4K1.
authors.forename: Teresita M
authors.initials: TM
authors.lastname: Porter
authors.affiliation: Department of Integrative Biology and Biodiversity Institute of Ontario, University of Guelph, 50 Stone Road East, Guelph, ON, Canada N1G 2W1.
authors.forename: Joel F
authors.initials: JF
authors.lastname: Gibson
authors.affiliation: Department of Integrative Biology and Biodiversity Institute of Ontario, University of Guelph, 50 Stone Road East, Guelph, ON, Canada N1G 2W1.
authors.forename: Rafal
authors.initials: R
authors.lastname: Dobosz
authors.affiliation: Department of Biology, University of Pennsylvania, Philadelphia, PA, USA 19104.
authors.forename: Daniel H
authors.initials: DH
authors.lastname: Janzen
authors.affiliation: Department of Biology, University of Pennsylvania, Philadelphia, PA, USA 19104.
authors.forename: Winnie
authors.initials: W
authors.lastname: Hallwachs
authors.affiliation: Department of Biology, McMaster University, 1280 Main Street West, Hamilton, ON, Canada L8S 4K1.
authors.forename: G Brian
authors.initials: GB
authors.lastname: Golding
authors.affiliation: Department of Integrative Biology and Biodiversity Institute of Ontario, University of Guelph, 50 Stone Road East, Guelph, ON, Canada N1G 2W1.
authors.forename: Mehrdad
authors.initials: M
authors.lastname: Hajibabaei
mesh_terms: D000818:Animals; D001181:Arthropods / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D004247:DNA / Q000032:analysis*; D003576:Electron Transport Complex IV / Q000235:genetics; D059014:High-Throughput Nucleotide Sequencing; D008969:Molecular Sequence Data; D017422:Sequence Analysis, DNA
chemical_list: D004247:DNA; D003576:Electron Transport Complex IV