DNA Reaction - Partial Digestion with 4 bp Site Restriction Endonucleases


Jeff Lawrence

1. Dilute restriction endonuclease to 0.5 Unit/ml. Make 5 mL total.

2. Serially dilute 2 mL enzyme with 2 mL 1X reaction buffer. Create 0.25, 0.125, and 0.0625 Unit/ml dilutions.

3. Mix the following:
1 mL DNA (1 mg ) 1 mL 10X Buffer 2 mL Enzyme Dilution (either 1, 0.5, 0.25, or 0.125 Units) 7 mL Sterile Water 10 mL Total Volume 4. Incubate at the appropriate temperature for 15 minutes.

5. Kill reaction by adding 10 mL 10 mM EDTA and incubate at 65° for 10 min.


Last Update: Thursday, 19-Jun-2014 11:50:26 PDT
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Eric Kofoid eckofoid at ucdavis.edu