IGEM:IMPERIAL/2006/Protocols/mini prep of culture: Difference between revisions
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Miniprep of cultures | ===Miniprep of cultures=== | ||
* | *Transfer 1.5ml of culture into eppendorf tubes. | ||
*Spin for 20seconds and remove supernatant. | *Spin for 20seconds and remove supernatant. | ||
*Take out Solution I and III from the fridge with syringes. Solution II (if it is cold it precipitates) is on the bench. (use yellow pipette for syringes) | *Take out Solution I and III from the fridge with syringes. Solution II (if it is cold it precipitates) is on the bench. (use yellow pipette for syringes) | ||
Line 11: | Line 11: | ||
*Pour the supernatants in to the new labelled tubes, add 1ml of ethanol and mix. | *Pour the supernatants in to the new labelled tubes, add 1ml of ethanol and mix. | ||
*Spin for another 3min and remove supernatant. | *Spin for another 3min and remove supernatant. | ||
*Spin to remove residual liquid and then resuspend in 50ul of MiliQ ( | *Spin to remove residual liquid and then resuspend in 50ul of MiliQ (1 sec. vortex. Don’t resuspend the pellet). | ||
vortex. Don’t resuspend the pellet). | |||
*NB: If the culture was grown during the day (8hrs), then resuspend in 25{{uL}} to obtain approximately the same concentration. | *NB: If the culture was grown during the day (8hrs), then resuspend in 25{{uL}} to obtain approximately the same concentration. | ||
*Spin for 1min. | *Spin for 1min. |
Revision as of 07:00, 28 October 2006
Miniprep of cultures
- Transfer 1.5ml of culture into eppendorf tubes.
- Spin for 20seconds and remove supernatant.
- Take out Solution I and III from the fridge with syringes. Solution II (if it is cold it precipitates) is on the bench. (use yellow pipette for syringes)
- Add 100ul of solution I and resuspend the pellet by vortexing.
- Add 200ul of solution II.
- Add 150ul of solution III.
- Mix them well by inverting the tubes. White precipitates appears.
- Spin for 3min. In the meantime, label tubes.
- Pour the supernatants in to the new labelled tubes, add 1ml of ethanol and mix.
- Spin for another 3min and remove supernatant.
- Spin to remove residual liquid and then resuspend in 50ul of MiliQ (1 sec. vortex. Don’t resuspend the pellet).
- NB: If the culture was grown during the day (8hrs), then resuspend in 25μL to obtain approximately the same concentration.
- Spin for 1min.
- Ready to digest.
Solution 1
- 90g glucose
- 10ml 0.5M EDTA
- 6.25ml 2M Tris pH8
- make upto 500ml
Solution 2
- For 500 mls
- 50ml 10%SDS
- 20ml NaOH
Solution 3
- 147g potassium acetate
- 57.5ml glacial acetic acid
- make upto 500ml