DNA Barcoding

The New York Botanical Garden Forest:

A Floristic Approach

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 Laboratory following universally agreed upon protocols for DNA Barcoding as proposed by CBOL and the Plant Working Group.

 

 

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For more information contact: Dr. Ken Cameron

 

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