Defining a DNA barcording locus for protists

Last month's PLoS Biology had a community article devoted to the Protist Working Group recently initiated though the Consortium for the Barcode of Life (CBOL). Now, people have mixed (often vehement) opinions about CBOL - I won't go into the history or debate about DNA barcoding here, but it's worth checking out posts by Dave Lunt at EvoPhylo (Rewriting the invention of DNA barcoding) and Jonathan Eisen at the Tree of Life ("Barcoding" researchers keep ignoring microbes and history).

Since its inception, CBOL has been overwhelmingly focused on the mitochondrial Cytochrome c Oxidase 1 loci that can be "universally" amplified (unless you're trying to barcode nematodes or any other taxon where these universal primer sets don't work). However, in recent years I've been pleased to see the formation of different taxon-specific working groups focused on ribosomal rRNA genes as "official" DNA barcodes (e.g. ITS for Fungi). Ribosomal loci have a much longer history--and thus more available reference data--in most eukaryote groups, and have pretty much been adopted as de facto barcodes for molecular studies (read: not yet CBOL-approved).

Even if all researchers working on a specific taxon are using the same gene to complement morphological/ecological information, I still strongly support the formation of these CBOL working groups. As more people adopt high-throughput sequencing approaches, we need coordination and interaction across different taxonomic communities. For a given taxon, discussions focused on the barcoding locus will simply get people talking, illuminating what different labs are actually doing and helping us to determine the most useful (although probably not perfect) community standards.

So CBOL working groups essentially gather all the experts within a particular taxon, and have them discuss the merits and drawbacks of different loci for molecular identification of species:

Identifying the standard barcode regions for protists and assembling a reference library are the main objectives of the Protist Working Group (ProWG), initiated by the Consortium for the Barcode of Life (CBOL, http://www.barcodeoflife.org/). The ProWG unites a panel of international experts in protist taxonomy and ecology, with the aim to assess and unify the efforts to identify the barcode regions across all protist lineages, create an integrated plan to finalize the selection, and launch projects that would populate the reference barcode library. (Pawlowski et al. 2012)

I was highly encouraged by the Protist Working Group's stance - instead of trying to force everyone to use a single locus (e.g. defining ONE barcode for all protists), they advocate a much more realistic approach:

Because of their long, independent, and complex evolutionary histories, protists are so genetically variable that it is virtually impossible to find a single universal DNA barcode suitable for all of them. The ProWG consortium therefore recommends a two-step barcoding approach, comprising a preliminary identification using a universal eukaryotic barcode, called the pre-barcode, followed by a species-level assignment using a group-specific barcode (Figure 3). In this nested strategy, the ~500 bp variable V4 region of 18S rDNA is proposed as the universal eukaryotic pre-barcode. Group-specific barcodes (Figure 2C) will then have to be defined separately for each major protistan group, based on comparative studies using the CBOL selection criteria, and much of this work is still to be done. (Pawlowski et al. 2012)

This proposed approach will easily translate to high-throughput studies - you might want to get a broad overview of eukaryote communities with a universal 18S primer set, and then dig deeper into species assemblages by also sequencing other loci (ITS, COX1) targeted at ecologically important groups.

Now all we need is a CBOL Working Group for Nematodes - seriously, why don't we have one yet?!

Reference:

Pawlowski J, Audic S, Adl S, Bass D, Belbahri L, Berney C, et al. (2012) CBOL Protist Working Group: Barcoding Eukaryotic Richness beyond the Animal, Plant, and Fungal Kingdoms. PLoS Biology, 10(11): e1001419.