Dear all
Sorry for off topic and lengthy post, but I came across a very unique DNA
contamination during one membrane protein purification (a microbial
external environment sensor/response protein)
Already done
* DNAse used as stranded protocol during cell break.
* Membrane extraction to
Dear Neeraj,
Do you mean the H3C protease ( trade name: PreScission protease provided by
GE)?
This protease can specifically cleavage the sequence encode by plasmid pGEX-6p,
which possess the GST tag.
Best wishes!
Lu Zuokun
--
卢作焜
南开大学新生物站A202
Lu Zuokun, Ph.D. Candidate
College of
Dear Pramod Kumar,
if the DNA binds to the protein, wouldn't this affect the interpretation
of the Agarose gel?
500 bases should result in a distinct shift during gel filtration. Do
you observe this?
While waiting for suggestions, you might set up crystallisation trials
just in case?
Best
pramod
Do a spin after bacterial cell breaking at 8K to get rid of a nuclear pellet
Also there is a nuclease preparation available called benzonase
it is most effective when you also add magnesium in your buffer
so in your homogenization buffer add magnesium and should avoid any EDTA
Pius
On Thu,
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Stepwise addition to 1% PEI (polyethylenimine) following cell lysis
(before dialysis) should do the trick.
--paul
On 06/25/2015 05:23 PM, Pramod Kumar wrote
Dear all
Sorry for off topic and lengthy post, but I came across a very unique
DNA contamination during one membrane protein
Dear all,
I am new to saxs and I get the model by saxs data and I have a
structure. I want to fit the structure to the shape but I can just open
them in PyMol.
Can any one teach me to fit them?
Best,
Weifei
Several different approaches may help you to separate DNA from protein
including;
1) When bound to the nickel column, wash the protein/beads with 1-2M NaCl or
1-2M KCl for a few hours to overnight. Increase ionic strength should disrupt
protein-DNA interactions.
2) Use a low concentration of
I think the more interesting questions are: should one want to disrupt such a
tight interaction?
Wouldn’t the structure of the protein bound to DNA be more interesting than the
protein alone?
Why not try to crystallize the complex and show how the protein binds?
Sometimes you should just run
We are currently looking for a highly motivated Ph.D. student for our
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Our work involves biochemical and structural studies on cellular
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