[Coral-List] DNA extraction and DNA barcoding without zooxanthellae DNA
zforsman2001 at yahoo.com.au
Thu Jun 7 19:43:40 EDT 2007
Dear Bill and Coral-listers,
I advocate using the ITS region for "barcoding" of zooanthids and scleractinians. mtDNA evolves very slowly and may not provide much resolution unless multiple genes are used (COI and CytB might be good bets when combined). The upside of ITS is that the primers are very conserved and almost always work, some primers are coral specific (but they don't always work). If you clone with universal primers, you can get both zoox and coral sequences in one shot -- the zoox sequences are always easy to determine by comparison to GenBank. The downside is that you may need to clone, because some groups have more than one ITS sequence type per genome (usually only in cases where hybridization events have occurred) -See Coleman (2003) for review. It is becoming clear that intragenomic variation is an exception and not the rule in corals (Wei et al. 2006).
When you blast an ITS seqeunce against the GenBank database, the results usually come back nearly in phylogenetic order. The ITS region holds a great deal of information that has yet to be fully exploited and in my opinion it will prove to be a more universal "barcode" for Eukaryotes.
Zac Forsman, Ph.D.
Zac at hawaii.edu
University of Hawaii
Dept. of Biology
2450 Campus Road, Dean II Bldg
Honolulu Hawai'i 96822
Coleman, AW (2003) ITS2 is a
double-edged tool for eukaryote evolutionary comparisons. Trends Genet. 19:
Alvarez I, Wendel JF (2003)
Ribosomal ITS sequences and plant phylogenetic inference. Mol Phyl Evol 29:
The ITS region in corals:
Wei NV, Wallace CC, Dai CF, Pillay
KRM4, Chen CA (2006) Analyses of the Ribosomal Internal Transcribed Spacers
(ITS) and the 5.8S Gene Indicate that Extremely High rDNA Heterogeneity is a
Unique Feature in the Scleractinian Cora Genus Acropora (Scleractinia;Acroporidae).
Zoological Studies 45(3):404-418
Forsman ZH (2003) Phylogeny
and Phylogeography of Porites and Siderastrea (Scleractinia: Cnidaria)
Species in The Caribbean and Eastern Pacific; Based on The Nuclear Ribosomal
ITS Region. Ph.D. Dissertation. University
Forsman ZH, Chen CA,
Fox GE, Wellington GM (2005) An ITS region phylogeny of Siderastrea (Cnidaria:Anthozoa): is S.glynni endangered or introduced? Coral
Forsman ZH, Wellington GM, Hunter CL, Fox GE (2006) Is
The ITS Region The Solution to The ‘Species Problem’ in corals? Intragenomic
Variation, and Alignment Permutation in Porites, Siderastrea and Outgroup Taxa. . 10th Int. Coral Reef Symp.14-23
S, Palumbi SR (2004) Testing the utility of ITS sequences in coral. Phylo Mol Ecol 13:2763-2772
J. D. Reimer · K. Takishita · S. Ono · T. Maruyama (in press) Diversity and evolution in the zoanthid genus Palythoa (Cnidaria: Hexacorallia) based on nuclear ITS-rDNA
Coral Reefs DOI 10.1007/s00338-007-0210-5 123
PS, I am finding new Porites species with ITS, and the results are congruent with COI with greater resolution (unpublished data). Please don't hesitate to contact me for references or primer sequences.
----- Original Message ----
From: "capman at augsburg.edu" <capman at augsburg.edu>
To: coral-list at coral.aoml.noaa.gov
Sent: Wednesday, June 6, 2007 11:02:02 PM
Subject: [Coral-List] DNA extraction and DNA barcoding without zooxanthellae DNA
One of my students is interested in trying to do DNA barcoding of
zoanthids and possibly some other cnidarians.
We have been researching DNA extraction methods that avoid DNA from
the zooxanthellae. It is seeming that one approach that has been
used is to extract DNA from pedal disk tissue. However, with some
species isolating tissue with no zooxanthellae is seeming to be
easier said than done, and most of the cnidarian DNA extraction
methods we have encountered have been somewhat vague in spots.
We would very much like to find someone experienced in these
techniques who would be willing to discuss methodologies with us by
phone or e-mail.
Also, it appears that the standard gene used for DNA barcoding of
animals is not suitable for Cnidaria. If any of you are successfully
barcoding cnidarians, we would be very interested in talking to you
If you are knowledgeable in these areas, or if you can suggest
someone who is, we would very much like to hear from you,
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