DNA Barcoding
The
A Floristic Ap
to Identifying a DNA Barcode for Plants
Project Materials and Methods
Zoologists have generally agreed
that a short, 600 base pair fragment of DNA from the mitochondrial genome (CO1)
exhibits sufficient variation to allow for unique identification of animal
species. Unfortunately, that region is
not sufficiently variable in plants to serve as a universal DNA barcode. A number of alternative gene regions have
been proposed [see Kress, W.J., K.J. Wurdack, E.A. Zimmer, L.A. Weigt, and D.H.
Janzen. 2005. Use of DNA 
barcodes to identify flowering plants. Proc. Natl. Acad. Sci. 102(23):8369-74 for a thorough discussion], but there has been
no consensus within the international botanical community as to which
single locus would serve most
effectively as a plant DNA barcode.
The goal of Phase I of the project
(completed in December 2005) was to identify three or more candidate gene
regions from a small set of plants that could be further tested across the
entire plant kingdom in Phase II of the project. There was consensus among the group that such
loci needed to meet the following criteria: 1) they should be located from
within the plastid genome, rather than nuclear or mitochondrial; 2) they should
be coding genes rather than intergenic spacers; 3) they should be of a total
length not exceeding 800 base pairs; 4) they should exhibit only minimal length
variation across the plant kingdom for alignment purposes; 5) they should be
amplified easily with universal primers;
and 6) they should exhibit sufficient variation to distinguish among
closely related species. During a
three-day workshop held at Kew Gardens from December 11-13, 2005, the Plant
Working Group's Principal Investigators were presented with the findings of
Phase I, which identified YCF5, accD, rpoB, and rpoC1 as
primary target loci best fulfilling these criteria, with matK and ndhJ as
secondary reserves. The results were based on a novel method of universal
amplification primer design devised by Drs. Michael Wilkinson and Nicola Toomey
at Reading University, followed by plastid genome sequencing of 122 plants
representing 61 phylogenetic sister-species pairs from across the entire plant
tree of life. [ See Project Rationale]
These same primary target gene
segments have been sequenced for the plant species targeted within the
Botanical Garden Forest. Each species
has been newly collected, identified by at least two staff botanists, and
pressed to serve as a new voucher in the Steere Herbarium. Newly collected leaf tissue has been
preserved in silica gel and frozen to serve as a source of total genomic DNA
for the project. All laboratory methods
are being carried out within the state-of-the-art facilities of the Botanical
Garden's new Pfizer Plant Research
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For more information
contact: Dr. Ken
Cameron
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