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Knight:Purification of His-tagged proteins/Denaturing: Difference between revisions
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Knight:Purification of His-tagged proteins/Denaturing (view source)
Revision as of 08:51, 5 October 2006
, 5 October 2006→Procedure
| Line 50: | Line 50: | ||
#Save 20 μL cleared lysate. | #Save 20 μL cleared lysate. | ||
#Load 600 μL cleared lysate to Ni-NTA column. | #Load 600 μL cleared lysate to Ni-NTA column. | ||
#Centrifuge Ni-NTA column | #Centrifuge Ni-NTA column 5 mins at 700 x ''g'' with closed lid. | ||
#*'' | #*''I typically repeat this step to load the rest of my cleared lysate.'' | ||
#*''Save flow through.'' | #*''Save flow through.'' | ||
#Add 600 μL wash buffer to Ni-NTA column. | #Add 600 μL wash buffer to Ni-NTA column. | ||
| Line 63: | Line 63: | ||
#Centrifuge Ni-NTA column 2 mins at 700 x ''g'' with open lid. | #Centrifuge Ni-NTA column 2 mins at 700 x ''g'' with open lid. | ||
#*''Most of the protein should elute in this elution step.'' | #*''Most of the protein should elute in this elution step.'' | ||
==Notes== | ==Notes== | ||
