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Team:UC Davis/Protocols
From 2011.igem.org
(Difference between revisions)
(Created page with "=Restriction Enzyme Double Digest= ====Materials==== *- 22 uL dH2O *- 1 uL BSA *- 5 uL Buffer *- 20 uL Template *- 1 uL Enzyme 1 *- 1 uL Enzyme 2 ====Buffer Compatibility Chart...") |
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====Materials==== | ====Materials==== | ||
| - | * | + | * 22 uL dH2O |
| - | * | + | * 1 uL BSA |
| - | * | + | * 5 uL Buffer |
| - | * | + | * 20 uL Template |
| - | * | + | * 1 uL Enzyme 1 |
| - | * | + | * 1 uL Enzyme 2 |
====Buffer Compatibility Chart==== | ====Buffer Compatibility Chart==== | ||
| Line 29: | Line 29: | ||
====Procedure==== | ====Procedure==== | ||
| - | * | + | * Mix reactants thoroughly. |
| - | * | + | * Place at 37 C for 3 hours. |
| - | * | + | * Increase to 80 C for 20 minutes to kill enzymes (some enzymes need only a 65 C heatkill, check enzyme). |
| - | * | + | * Run on a gel and extract product. |
| - | + | ||
=Gel Extraction/Purification Procedure= | =Gel Extraction/Purification Procedure= | ||
Revision as of 18:05, 28 June 2011
Contents |
Restriction Enzyme Double Digest
Materials
- 22 uL dH2O
- 1 uL BSA
- 5 uL Buffer
- 20 uL Template
- 1 uL Enzyme 1
- 1 uL Enzyme 2
Buffer Compatibility Chart
| 1 | 2 | 3 | 4 | |
|---|---|---|---|---|
| EcoRI | 100 | 100 | 100 | 100 |
| SpeI | 75 | 100 | 25 | 100 |
| PstI | 75 | 75 | 100 | 50 |
| NheI | 100 | 100 | 10 | 100 |
| XbaI | 0 | 100 | 75 | 100 |
Procedure
- Mix reactants thoroughly.
- Place at 37 C for 3 hours.
- Increase to 80 C for 20 minutes to kill enzymes (some enzymes need only a 65 C heatkill, check enzyme).
- Run on a gel and extract product.
