Case Study
Title & Categories
| Case Study Title: | Barcoding Darwin’s meadow: a 21st century botanical inventory at historic Down House |
| Focus Theme: | Resolving discrete taxonomic problems, Adding barcodes to a large survey of a taxonomic group, Invasive species, Systematic revision of an entire taxon, Associating life stages/genders, Agricultural pests, Resolving discrete taxonomic problems |
| Geographic Region: | Southern Asia, Oceania, South East Asian Peninsula, Unspecified |
| Habitat Type: | Freshwater rivers, Freshwater rivers, Terrestrial temperate forest, Terrestrial grasslands, Terrestrial cultivated land, Marine - Other |
| Taxonomic Group: | f: VERTEBRATES, f: Osteichthyes, f: Chondrichthyes, d: Arthropoda, d: Arthropoda - Hexapoda - Other Insecta, d: Arthropoda - Hexapoda - Lepidoptera, d: Arthropoda - Hexapoda - Diptera, d: Cnidaria |
| Project Duration: | 5/2007 to 12/2007 |
Scope
We have collected ~200 specimens of ~160 flowering plant species with 5-10 fold sampling density for ~1-5 species. We will sequence all four loci proposed by Chase et al. (2007), that is, rpoC1, rpoB, matK, trnH-psbA, as well as rbcL-a as proposed by Kress et al. (2007).
Purpose
To survey Great Pucklands meadow at Down House, Kent with the following specific aims:
1) develop a seamless approach to collection, management and storage of morphological and molecular collections for the 21st century including high throughput DNA barcode sequencing
2) measure amplification and discrimination performance of the proposed combinatorial plant barcodes (including rpoC, rpoB, matK, trnH-psbA and rbcL
3) carry out a controlled test of Whatman FTA sample collection and DNA isolation technology vs. our existing method of silica desiccation followed by CTAB extraction
Background
In 1855, Charles Darwin surveyed Great Pucklands meadow near his home at Down House. In an age when specimen rarity was prized above all, Darwin’s vision was radical: to identify all plant species in a small, unremarkable plot. As it did for Darwin, Great Pucklands still serves as an ideal training ground for the development of plant identification skills.
Logistics
In this project we are piloting procedures for the development of future Natural History Museum botanical collections and analysis. These include:
1) coordination of specimen data using the NHM's new KE database, including morphological voucher specimens, DNA sample storage locations/parameters and multi-locus sequence data/ alignments
2) collection, storage and utilisation of DNA specimens on Whatman FTA cards vs. standard method of silica-desiccation or -20 degree storage of DNA extracts
3) high-throughput DNA sequencing procedures involving automated cellular disruption in 8-tube strips and DNA extraction in 96-tube racks (12 strips of 8), followed by 96-well format PCR, purification and quantification, visualisation using E-Gel-96 system from Invitrogen, and SeqScape vs. MBToolbox software for processing 96-sample batches of sequence data