Protocols

The following protocols are developed for the Research and Conservation Department at Denver Botanic Gardens, but may be used as general guidelines for similar research projects or programs.

This section is a work in progress. Please return periodically to find more protocols and instructions in everything from collecting specimens in the field, to DNA extraction and sequencing.

Field Protocols

Herbarium Protocols

Laboratory Protocols

  • DNA Extraction
  • PCR protocol – For our protocol we use the Bioline MyTaq mix, which works well.
    • Primer page for info on Sanger and High Throughput Sequencing (HTS). (Coming soon[ish])
  • Gel Electrophoresis – Pretty standard (Coming soon[ish])
    • Pouring Gels
    • TBE buffer instructions
  • PCR Clean and DNA quantification – We use a simple two step alcohol precipitation to clean PCR product. For DNA quantification we have a NanoDrop (not recommended for precise measurements) and a Qubit.
    • Cycle sequencing clean – The Field Museum method. Basically an alcohol precip method but with EDTH buffer. (Coming soon[ish])
  • Sanger Sequencing Shipping Instructions (Coming soon[ish])
  • High Throughput Shipping Instructions (Coming soon[ish])

Data Analysis

  • Editing ABI files in Geneious (Coming soon[ish])
  • Assembling Datasets and Performing Phylogenetic Analyses – This process is pretty convoluted. It requires a number of steps that can easily get lost in translation in a document like this. Hopefully someday we will have funding to support workshops that detail this process.