User:Behzad Damadzadeh/Notebook/PcTF Subcloning in E-coli/2012/05/23: Difference between revisions
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Note that prestained molecular weight marker doesn't need any preparation. | Note that prestained molecular weight marker doesn't need any preparation. | ||
After the heating process keep the samples on the bench to cool down to room temperature. | After the heating process keep the samples on the bench to cool down to the room temperature. | ||
===Running the gel=== | ===Running the gel=== | ||
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#Fill the upper buffer chamber with a small amount of upper buffer chamber running buffer (with antioxidant) to check tightness of seal. | #Fill the upper buffer chamber with a small amount of upper buffer chamber running buffer (with antioxidant) to check tightness of seal. | ||
#*If there is a leak, discard buffer, reseal chamber and try again. | #*If there is a leak, discard buffer, reseal chamber and try again. | ||
#Fill upper buffer chamber. Buffer level should exceed level of the wells. Requires about | #Fill upper buffer chamber. Buffer level should exceed level of the wells. Requires about 400mL | ||
#Load samples. | #Load samples. | ||
#Load protein molecular weight marker (20 μL per lane but 10μL also seems to work). | #Load protein molecular weight marker (20 μL per lane but 10μL also seems to work). | ||
#Fill lower buffer chamber at the gap near locking mechanism with | #Fill lower buffer chamber at the gap near locking mechanism until the upper part of the removed white tape with 100mL NuPAGE SDS running buffer. | ||
#Run at | #Run at 120V until the samples running down to the nearest bottom edge of the gel. | ||
===Staining the gel=== | ===Staining the gel=== |
Revision as of 18:08, 24 May 2012
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5/23/2012Materials
ProcedureRunning buffer preparationDo this step first.
Sample preparation
Set up the gel apparatus during the 10 mins sample heating step. Note that prestained molecular weight marker doesn't need any preparation. After the heating process keep the samples on the bench to cool down to the room temperature. Running the gel
Staining the gel
You have three options for staining at this point.
See Knight:NuPAGE electrophoresis/Gel drying for instructions on how to dry and preserve the gel.
Marker Sizes
Notes
Safety
See here for detailed safety information. References |