DNA Extraction From Fungal Plate
Please record all extractions correctly in the database. See database protocols for isolations for more information. Below is the current method:
General Ex-N-Amp protocol:
- Prepare strip tubes with 40uL of Sigma Aldrich extraction solution. This can be found in aliquots in the shared box in the door of the small freezer.
- Scrape approximately 10uL of hyphae from fungal colony using a sterile pipette tip (20uL works best and is not used for much else) or scalpel.
- Add fungal material to extraction solution and vortex.
- Run Ex-N-A protocol on a thermocycler (96C for 30 minutes).
- Spin down with high rpm.
- Pipet off the top 25uL: that’s your extract. It is often safe to dilute this a bit if you will need more than 25 uL, but this is not standard procedure.
Note: If BSA is needed, we have a stock of 2% molecular grade BSA in our freezer. This should be made into aliquots before using for samples. This is not standard, and only used in cases where it increases DNA amplification success.
OLD: DNA Extraction from mycangia
Modified Sigma Ex-N-Amp protocol:
- add contents of mycangium (if dissectable) or the body part with mycangium (for example, mandibular mycangium) to 10uL of XNA Extraction solution in a PCR tube
- mash the sample with a melted pipette tip
- run Ex-N-A protocol on a thermocycler (96C for 10 minutes)
- add 10uL of 3% BSA. DON’T ADD THE STANDARD DILUTION SOLUTION FROM SIGMA! It decreases yield sugnificantly.
- spin down with high rpm
- pipet off the top 10uL, and dilute with 10uL of water: that’s your extract