Dear All,
I am having problems in extracting the transmembrane proteins from Symbiodinium, but no problems with cytoplasmic proteins. The extraction medium I am using consists of Tris, sodium deoxycholate, NP-40, NaCl, EDTA and roche protease inhibitors. Then I diluted the homogenate in Laemmli buffer and no heating is applied to the sample. When I loaded 100ug, I could not detect the band of interest for all of the transmembrane proteins.
I have used this extraction buffer for animal transmembrane proteins and it has worked pretty well but not for the Symbiodinium.
Can anyone help me with this? Thanks.
Hi Mel, you could try triton X-114 phase separation.
Article Triton X-114 phase separation in the isolation and purificat...
The reason you are not seeing your protein on a gel might be because the sample already has lots of detergents in it and the SDS in the sample buffer is not able to exert its effects.
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