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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 07:51, 22 September 2006
, 22 September 2006→Procedure
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#*''Since I don't know what yield to expect, I arbitrarily did 50mL cultures assuming that my protein yield would be low and let it grow most of the day. From looking at the culture, it was near but not at saturation but I didn't have time to take OD readings.'' | #*''Since I don't know what yield to expect, I arbitrarily did 50mL cultures assuming that my protein yield would be low and let it grow most of the day. From looking at the culture, it was near but not at saturation but I didn't have time to take OD readings.'' | ||
#[[Knight:pH meter/Measurement|Verify pH]] of lysis, wash and elution buffers. Adjust if necessary. | #[[Knight:pH meter/Measurement|Verify pH]] of lysis, wash and elution buffers. Adjust if necessary. | ||
#*''Dissociation of urea can lead to changes in pH.'' | #*''Dissociation of urea can lead to changes in pH. The pH definitely needs to be checked prior to using the solutions.'' | ||
#Harvest the cells by centrifugation at 4000 x ''g'' for 15 mins. | #Harvest the cells by centrifugation at 4000 x ''g'' for 15 mins. | ||
#*''The Qiagen protocol didn't specify a temperature so I did 4°C.'' | #*''The Qiagen protocol didn't specify a temperature so I did 4°C.'' | ||
