Streamlining DNA barcoding protocols: automated DNA extraction and a new cox1 primer in arachnid systematics

PloS One
Nina VidergarMatjaž Kuntner

Abstract

DNA barcoding is a popular tool in taxonomic and phylogenetic studies, but for most animal lineages protocols for obtaining the barcoding sequences--mitochondrial cytochrome C oxidase subunit I (cox1 AKA CO1)--are not standardized. Our aim was to explore an optimal strategy for arachnids, focusing on the species-richest lineage, spiders by (1) improving an automated DNA extraction protocol, (2) testing the performance of commonly used primer combinations, and (3) developing a new cox1 primer suitable for more efficient alignment and phylogenetic analyses. We used exemplars of 15 species from all major spider clades, processed a range of spider tissues of varying size and quality, optimized genomic DNA extraction using the MagMAX Express magnetic particle processor-an automated high throughput DNA extraction system-and tested cox1 amplification protocols emphasizing the standard barcoding region using ten routinely employed primer pairs. The best results were obtained with the commonly used Folmer primers (LCO1490/HCO2198) that capture the standard barcode region, and with the C1-J-2183/C1-N-2776 primer pair that amplifies its extension. However, C1-J-2183 is designed too close to HCO2198 for well-interpreted, continuous sequenc...Continue Reading

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Citations

Aug 23, 2016·PeerJ·Jonathan A CoddingtonMatjaž Kuntner
Nov 15, 2017·Parasites & Vectors·Nídia CangiNathalie Vachiéry
Jan 25, 2019·Scientific Reports·Klemen ČandekMatjaž Kuntner
Sep 2, 2017·Ecology and Evolution·Corinna WallingerMichael Traugott

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Methods Mentioned

BETA
PCR
Electrophoresis
PCRs

Software Mentioned

Primer3
Mesquite
Geneious Pro

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