DNA Extraction protocols

From Tia, dated 15 DEC 2016: Protocol modifications with Qiagen DNeasy Mini Plant Kit

Species: Rumex rothschildianus

• Pick young, small, green leaves to fill tubes
• Use 3-4 leaves per tube
• Use 2 medium sized glass beads
• “Pre-crush” frozen tissue with pipette tips before using the Tissue Lyser
• Frozen tubes can be stored in the -80°C freezer for months before extraction
• After the second wash step with buffer AW2 perform 1 dry centrifuge (1min at 8000rpm) to dry surface of filter paper (add no buffer)
• Perform 2 incubation steps, 10 minutes each, 50 ul of sterile water per incubation (to give a total of 70ul DNA sample)

Species: Rumex hastatulus

• Pick young, small, green leaves to fill tubes
• Use as many young leaves as you can fit in each tube
• Use 2 medium sized glass beads
• “Pre-crush” frozen tissue with pipette tips before using the Tissue Lyser
• Frozen tubes can be stored in the -80°C freezer for a few weeks before extraction
• Only perform 2-4 preps at a time to prevent thawing of sample during extraction process
• Perform 2 incubation steps, 10 minutes each, 35 ul of sterile water per incubation (to give a total of 70ul DNA sample)
• If DNA yields are low, perform multiple preps (4-6) for each sample, combine samples into one tube, incubate sample on 65°C to evaporate water and increase concentration

Species: Capsella grandiflora

• Pick young, small, green leaves to fill tubes
• Use 3-4 leaves per tube
• Use 2 medium sized glass beads
• Frozen tubes can be stored in the -80°C freezer for months before extraction
• Can use the DNA extraction robot for extractions or perform extractions by hand according to Qiagen instructions
• Perform 2 incubation steps, 5 minutes each, 50 ul of sterile water or elution buffer per incubation (to give a total of 100ul of DNA sample)