Lidstrom:Digestion with Restriction Enzymes: Difference between revisions
From OpenWetWare
Jump to navigationJump to search
m (→Recipe) |
(→Recipe) |
||
| Line 11: | Line 11: | ||
**If two enzymes are required, check http://www.neb.com/nebecomm/DoubleDigestCalculator.asp for buffer, temperature, BSA specifications. | **If two enzymes are required, check http://www.neb.com/nebecomm/DoubleDigestCalculator.asp for buffer, temperature, BSA specifications. | ||
*If using BioBrick or BglBrick enzymes, chose your buffer based on Janet's notes. | *If using BioBrick or BglBrick enzymes, chose your buffer based on [https://docs.google.com/document/d/1j91GWl_90ZablSxtE-hFQWPhG3BHQx822zSvSH1dv04/edit Janet's notes]. | ||
== Temperature/Time == | == Temperature/Time == | ||
*1 - 2 hours at 37oC or room temperature overnight | *1 - 2 hours at 37oC or room temperature overnight | ||
*Inactivate by .. | *Inactivate by .. | ||
Revision as of 08:33, 30 March 2013
Back to Protocols
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 ..
