Express barcodes: racing from specimen to identification

Abstract

Although devices combining microfluidic and advanced sequencing technologies promise a future where one can generate a DNA barcode in minutes, current analytical regimes typically involve workflows that extend over 2 days. Here we describe simple protocols enabling the advance from a specimen to barcode-based identification in less than 2 h. The protocols use frozen or lyophilized reagents that can be prepackaged into 'kits' and support barcode analysis across the animal kingdom. The analytical procedure allows 5 min for DNA extraction, 25 min for polymerase chain reaction amplification of the barcode region, 25 min for cycle-sequencing, 10 min for cleanup, 45 min for capillary sequencing and 5 min for trace file analysis to complete DNA-based identification. This study involved the comparison of varied DNA preservation and extraction methods, and evaluated Taq polymerases with high processivity and resistance to inhibitors.

Life history and DNA barcode of Oxyurella longicaudis (Birgei, 1910) (Cladocera, Anomopoda, Chydoridae)

“…To amplify the mitochondrial COI gene, the universal primers, LCO 1490 and HCO2198 (Folmer et al 1994) were used. PCR reactions had a total volume of 25 μl and were performed according to Ivanova et al (2009) using Platinum Taq (Invitrogen, Carlsbad, CA, USA) as the enzyme. The PCR conditions were 94°C for 2 min as initial denaturation and 40 cycles of 94°C for 40s, 55°C for 40s, and 72°C for 1 min.…”

DNA Barcodes for Marine Biodiversity: Moving Fast Forward?

“…Non-specialists are able to have a fast (express-barcoding in less than two hours [47]), cheap and reliable identification tool with many practical and fundamental applications. Moreover, there is an international Consortium for the Barcode of Life (CBOL; http://www.barcoding.si.edu) dedicated to establish DNA barcoding as a standard tool for species identification.…”

Mitochondrial pseudogenes in insect DNA barcoding: differing points of view on the same issue

“…DNA barcoding, a taxonomic method that uses a short, standardized DNA sequence to identify species, has gained increased attention and acceptance from members of the scientific community interested in documenting the Earths' biodiversity (Hebert et al 2003a, b, Savolainen et al 2005, Ivanova et al 2009, Janzen et al 2009). Since its inauguration in 2004, the Consortium for the Barcode of Life -CBOL, managed primarily by the Canadian Centre for DNA Barcoding at the Biodiversity Institute of Ontario, University of Guelph, Ontario, Canada, has gathered partners from all over the world.…”