I want to analyze the temperature mediated Chitinase/s production in Sugar beet tap root using2D electrophoresis. I prefer to retain the enzyme activity of the chitinase( Under Native conditions) Is it possible to run gels under Native conditions in 2 D electrophoresis.Thank you.
0
2 replies to this topic
#2
anonymous
-
-
- 1,890 posts
Veteran
0
Neutral
Neutral
Posted 09 September 2001 - 09:00 PM
Yes, you can but not by direct native gels. This might complicate your pattern completely, preventing analysis. Here is what we recommend. Run the first dimension ( the pI based separation ); run the second dimension under denaturing conditions but with two changes; reduce SDS content of the gel to 0.01% in the running buffer and the gel itself. Once the run is complete, soak the gel in 2% methanol at 37C for 2 hours. That should wake up your enzymes easily.
Good Luck
NikkiDr. Shankar's Technology Files, LLC[url="http://www.dstf.bigstep.com"][url]http://www.dstf.bigstep.com[/url][/url]--------------------------------------------
#3
anonymous
-
-
- 1,890 posts
Veteran
0
Neutral
Neutral
Posted 09 September 2001 - 09:00 PM
Yes, you can but not by direct native gels. This might complicate your pattern completely, preventing analysis. Here is what we recommend. Run the first dimension ( the pI based separation ); run the second dimension under denaturing conditions but with two changes; reduce SDS content of the gel to 0.01% in the running buffer and the gel itself. Once the run is complete, soak the gel in 2% methanol at 37C for 2 hours. That should wake up your enzymes easily.
Good Luck
NikkiDr. Shankar's Technology Files, LLC[url="http://www.dstf.bigstep.com"][url]http://www.dstf.bigstep.com[/url][/url]--------------------------------------------
