Lidstrom:Digestion with Restriction Enzymes: Difference between revisions
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== Recipe == | == Recipe == | ||
*Try it quick and imprecise like [http://2011.igem.org/Team:Washington/Protocols/Digestion iGEM 2011 UW] recommends: | *Try it quick and imprecise like [http://2011.igem.org/Team:Washington/Protocols/Digestion iGEM 2011 UW] recommends: | ||
**10 uL DNA ( | **10+ uL DNA ( | ||
*** Want at least 500 ng. Can elute a miniprep in 38 uL of water and use that whole thing. Don't use miniprep DNA eluted in buffer; only use DNA eluted in water. | |||
**5uL buffer (check http://www.neb.com/nebecomm/EnzymeFinderSearchByName.asp for a list of enzymes and associated buffers) | **5uL buffer (check http://www.neb.com/nebecomm/EnzymeFinderSearchByName.asp for a list of enzymes and associated buffers) | ||
**0.5 uL BSA (if required. See above) | **0.5 uL BSA (if required. See above) |
Revision as of 15:04, 2 October 2012
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Recipe
- Try it quick and imprecise like iGEM 2011 UW recommends:
- 10+ uL DNA (
- Want at least 500 ng. Can elute a miniprep in 38 uL of water and use that whole thing. Don't use miniprep DNA eluted in buffer; only use DNA eluted in water.
- 5uL buffer (check http://www.neb.com/nebecomm/EnzymeFinderSearchByName.asp for a list of enzymes and associated buffers)
- 0.5 uL BSA (if required. See above)
- 1uL enzyme 1
- water to 50 uL (32.5 uL, add first)
- If two enzymes are required, check http://www.neb.com/nebecomm/DoubleDigestCalculator.asp for buffer, temperature, BSA specifications.
- 10+ uL DNA (
- If using BioBrick or BglBrick enzymes, chose your buffer based on Janet's notes.
Temperature/Time
- 1 - 2 hours at 37oC or room temperature overnight
- Inactivate by ..