Team:Brown/Notebook/Protocols
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#When complete, pour TBE buffer back into a container. The buffer can be used up to 8 times. | #When complete, pour TBE buffer back into a container. The buffer can be used up to 8 times. | ||
- | + | ===Ligation Reaction Procedure=== | |
#Thaw T4 DNA ligase and 10x Buffer on ice. | #Thaw T4 DNA ligase and 10x Buffer on ice. | ||
#In a sterile tube add (in this specific order): '''1.''' 8 microliters dd water '''2.''' 1 microliter vector '''3.''' 3 microliters insert '''4.''' 1.5 microliter 10x Ligase Buffer '''5.''' 1.5 microliter T4 DNA ligase | #In a sterile tube add (in this specific order): '''1.''' 8 microliters dd water '''2.''' 1 microliter vector '''3.''' 3 microliters insert '''4.''' 1.5 microliter 10x Ligase Buffer '''5.''' 1.5 microliter T4 DNA ligase | ||
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*The following equation can be used: Insert mass in nanograms = 3 x [Insert length (bp) / Vector length (bp)] x Vector Mass in nanograms | *The following equation can be used: Insert mass in nanograms = 3 x [Insert length (bp) / Vector length (bp)] x Vector Mass in nanograms | ||
- | + | ===Transformation Procedure=== | |
#Place both in ice. The control in this experiment is a negative control because no DNA is added to the competent cells. | #Place both in ice. The control in this experiment is a negative control because no DNA is added to the competent cells. | ||
#Obtain 100 microliters of competent cells for each centrifuge tube. Competent cells must thaw before being used. | #Obtain 100 microliters of competent cells for each centrifuge tube. Competent cells must thaw before being used. |
Revision as of 14:44, 25 October 2008
ProtocolsMiniprep Prodedure
Gel Electrophoresis
Ligation Reaction Procedure
Transformation Procedure
Ethanol Precipitation
Gel Extraction
The contents of the tube should be yellow after incubation.
Calf CIP - Calf Intestinal Alkaline Phosphatase
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