Naegleria 5
Naegleria 5 kdorfman Tue, 02/08/2022 - 21:088 QIAprep Spin miniprep kits for 4 cultures each
Notes before starting
- Optional: Add LyseBlue reagent to Buffer P1 at a ratio of 1 to 1000.
- Add the provided RNase A solution to Buffer P1, mix and store at 2–8°C.
- Add ethanol (96–100%) to Buffer PE before use (see bottle label for volume).
- All centrifugation steps are carried out at 13,000 rpm (~17,900 x g) in a conventional table-top microcentrifuge.
Each kit contains:
- 4 spin columns
- 6 buffers, as below
| Buffer | uL per rxn | mL per 4.5 rxns |
|---|---|---|
| P1 | 250 | 1.125 |
| P2 | 250 | 1.125 |
| N3 | 350 | 1.575 |
| PB | 500 | 2.25 |
| PE | 750 | 3.375 |
| EB | 50 | 0.225 |
Other materials
- Age1 + cutsmart
- 8 2mL tubes of LB broth
- competent HT115 cells from -80
- two 1% gels for wide gel rig, 2 small-well combs each
- 1X TAE running buffer
- 1KB DNA standard
Equipment
- Microcentrifuges capable of 13000rpm
- Hot blocks at
- 37
- 42
- two wide gel rigs
- two power supplies
- two blue light boxes with photography box