TITLE: DNA Barcoding Standard Operating Protocol, Plants and Lichens at RBGE, Sample Data
AUTHORS: Laura L Forrest, David Bell, Michelle Hart

[DESCRIPTION]
This is part of the collection DToL Taxon-specific Standard Operating Procedures for the Plant Working Group (protocols.io). The SOP collection contains guidance on how to process the various land plant taxa within the scope of the Darwin Tree of Life project. The guidance specifically refers to the metadata associated with tissue samples for DNA barcoding (which takes place at the Royal Botanic Garden (RBGE)). Every specimen is submitted for DNA barcoding first before potentially being sent to the Wellcome Sanger institute for whole genome sequencing.

This DNA barcoding SOP outlines the processing of plant and lichen sample data for the Darwin Tree of Life project (DToL) at the Royal Botanic Garden Edinburgh (RBGE). 

DNA barcoding is used as part of the species identification process AND sample tracking (to check that the genome sequence corresponds to the material that was sent and that there have been no sample mix-ups).

Definition: Land plants (Embryophyta) and lichens
Including: Bryophyta, Marchantiophyta, Anthocerotophyta, Lycopodiophyta, Polypodiophyta, Pinophyta, Cycadophyta, Ginkgophyta, Gnetophyta, Magnoliophyta, lichenized fungi

Excluding: all non-lichenized fungi

[GUIDELINES]

Including: Bryophyta, Marchantiophyta, Anthocerotophyta, Lycopodiophyta, Polypodiophyta, Pinophyta, Cycadophyta, Ginkgophyta, Gnetophyta, Magnoliophyta, lichenized fungi


Sample Tissue
Plant material (typically leaf or thallus material, but may be other plant tissue such as flowers) or lichen thallus tissue is collected and submitted to the Barcoding Centre following Standard Operation Protocols in the collection DToL Taxon-specific Standard Operating Procedures for the Plant Working Group (protocols.io) 


 


Required sample metadata
The collector or Genome Acquisition Lab emails a completed DToL SAMPLE MANIFEST for the collections that are to be barcoded to the Barcoding centre. This is then adapted to include one line per collection number, with all the Genome Acquisition Lab numbers (e.g. EDTOL or KDTOL numbers) that apply to that collection concatenated into a single cell (e.g. EDTOL00053,EDTOL00054,EDTOL00055). 

The data in this manifest is then used to produce:

1. EDNA numbers: These are in-house accession numbers that are given to RBGE DNA extractions. 
 



2. BOLD specimens (with Sample IDs and Process IDs): this is the data about each DToL DNA barcoding collection that is stored in the open access Barcode of Life database. Each specimen contains sample metadata, and can include image files (e.g. habitat photos, close-ups of fronds, flowers, leaves etc.) and a collection map.
 

For DToL barocding at RBGE, all the relevant collector, EDNA and BOLD numbers for each sample are pasted into a shared google sheet, RBGE DToL Barcoding. This sheet must NOT be sorted, as it contains several linked sheets. (To sort, copy and paste the relevant columns into a new spreadsheet).




 



 
     Version      Date      Major changes      Contributors        1.0      2020-04-07      Draft   version      Lyndall   Pereira (NHM)        2.0      2020-04-29      DNA   barcoding for reference library at NHM edited      NHM: Olga   Sivell, Laura Sivess, Gavin Broad, Ben Price, Lyndall Pereira        3.0      2020-05-13      Addition of   pictorial dissection for each taxon group      Liam   Crowley (Oxford WW) and Lyndall Pereira (NHM)        3.1      2020-07-17       SOP text   split into Order-specific SOPs. Uniform style, publication quality images   with colour blind friendly dissection lines      Olga   Sivell, Laura Sivess        4.0      2022-01-04      Overall   formatting further improved (including spelling/ punctuation changes, table   of contents links), photo guides made more uniform, edits in protocol made to   reflect any adaptations in the sampling process since July 2020, additional   taxon protocols included, unrequired taxon guides removed. First submission   to protocols.io.      Inez   Januszczak (NHM), Liam Crowley (Oxford WW), Olga Sivell (NHM), Laura Sivess   (NHM), Chris Fletcher (NHM)   

 

 Working SOP, checked by experts
 

[STEPS]