Haynes Lab:Notebook/Engineering PC-TFs/2013/03/18: Difference between revisions
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= 30.0 uL </font> | = 30.0 uL </font> | ||
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''Digest Order:'' | ''Digest Order:'' | ||
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*4 BL12 XbaI/PstI | *4 BL12 XbaI/PstI | ||
Follow steps for gel electrophoresis. | |||
* Orient top notch of gel electrophoresis apparatus. Place gel tray into larger tray. | |||
* Fill gel flask with up to 60 ml of TA buffer. | |||
* Create 1% gel by putting .6 grams of agarose into flask. | |||
* Microwave agarose solution for 40 seconds | |||
* Using hot gloves, grab flask out of microwave and swirl flask vigorously. Place back in microwave for 40 more seconds. | |||
* When flask is taken out of microwave, make sure that the agarose is completed dissolved. | |||
* Set up wells on gel electrophoresis apparatus. The mold for the first digest is B-10. Use the thick "teeth" and slip in the notches. | |||
* Add EtBr (ethidium bromide) to the agarose gel. There should be 1 microliter of EtBr per 10ml of agarose gel. Therefore, 6 µL. | |||
* Pour gel into tray. | |||
* Wash the agarose gel flask. | |||
* Run gel for 45 min at 100 V. Check gel under UV light. | |||
{| {{table}} cellspacing="3" <!-- Digest rxn. table --> | {| {{table}} cellspacing="3" <!-- Digest rxn. table --> |
Revision as of 20:30, 22 March 2013
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SummaryCutting inserts to be placed in mammalian vector
A) Get biobricks and enzymes from freezer. SpeI, PstI, XbaI The following list of parts are:
Digest Order: Biobrick Restriction Enzymes
Follow steps for gel electrophoresis.
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