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Hi all and a belated Happy New Year,
I was wondering if any of you noticed that the C6 Webtool from Janet Newman
(https://c6.csiro.au/C6.asp) is disco
ated cells if you are in a mammalian expression
system.
8. Try the same protein from other species
Janet Newman
Principal Scientist / Director, Collaborative Crystallisation Centre (C3)
CSIRO Material Science and Engineering
343 Royal Parade
Parkville. VIC. 3052
Australia
Tel +613 9662 7326
Email
Given that your tartrate crystals diffract so much better than your
non-tartrate crystals, and tartrate is found in the active site, you might
think that tartrate binding in the active site is related to the good
diffraction of your crystals. Soaking out a bound tartrate is going to be hard,
wh
?Hi Jonathan,
Hopefully you know about the trick of making any precious condition last longer
- use the 'magic' solution only in the drop itself, and use the best
approximation you can make in the reservoir of the experiment (if you are doing
vapour diffusion)
Regards, Janet
Ja
The way I have done it (and it was sort of fun in a mad scientist way) was to
mix up solid DL-malic acid and sodium hydroxide in the right amounts and add
water as needed. This generates of heat, but gets around the
solubility minimum which is impossible to get out of:
I mixed 40.23 g of DL-m
Just to point out that whatever buffer you purify your protein into is possibly
not the one that will keep your protein happiest. We had the opportunity of
testing about 250 proteins in DSF against 26 different buffer / salt
combinations (in triplicate, with lots of controls) and found out that
Hi All,
We have become quite interested in autoscoring of images of crystallization
droplets (I think everyone goes through this stage, most grow out of it
eventually) and as part of our work we wanted to develop a well-scored set of
images to use as a training set for machine learning approach
Generally one will make this up from stock solutions:
Make up
1 M Tris chloride pH 8
1 M magnesium chloride
50 % (weight/volume) PEG 6K
Then mix these together in the right ratios
(for 1 mL)
100 uL 1 M tris solution
200 uL 1 M MgCl2 solution
400 uL 50% PEG solution
300 uL water.
Janet
might be interested.
Regards, Janet
Dr Janet Newman
Principal Scientist | C3 Facility Manager
Materials Science and Engineering
CSIRO
Phone: +61 3 9662 7326 | Fax: +61 3 9662 7101 |
janet.new...@csiro.au | www.csiro.au | www.csiro.au/c3
Address: 343 Royal Parade, Parkville VIC 3052 AUSTRALIA
creen, you have to wonder (as Enrico points out) how
these can work, as the bits that change the most in any protein family are the
outside bits (ie, away from the active site) and thus are generally the parts
going to affect crystallisation the most.
Janet
Janet Newman
Principal Scientist
I would try 2.5M or 3M sodium malonate pH 7
Janet Newman
Principal Scientist / Director, Collaborative Crystallisation Centre
CSIRO Material Science and Engineering
343 Royal Parade
Parkville. VIC. 3052
Australia
Tel +613 9662 7326
Email janet.new...@csiro.au
We add 0.02% Azide to all the water used for washing in our Phoenix, and that
seems to help.
Janet Newman
Principal Scientist / Director, Collaborative Crystallisation Centre
CSIRO Material Science and Engineering
343 Royal Parade
Parkville. VIC. 3052
Australia
Tel +613 9662 7326
Email
round) since the screening.
Regards, Janet
Dr Janet Newman
Principal Scientist | C3 Facility Manager
Materials Science and Engineering
CSIRO
Phone: +61 3 9662 7326 | Fax: +61 3 9662 7101 |
janet.new...@csiro.au | www.csiro.au | www.csiro.au/c3
Address: 343 Royal Parade, Parkville VIC 3052 AUST
Hi Jin,
Could it be the wrong amino acid in the sequence? Have you sequenced the gene
construct that you used? The codon usage for HIS is CAC or CAU, and for TYR is
UAC or UAU. -
Cheers, Janet
From: CCP4 bulletin board [ccp...@jiscmail.ac.uk] On Be
One thing to do here is to work out where you have your problem.
Is the crystal mosaic after you have introduced your potential cryo solutions,
but still at room temperature? If it is, there is very little chance
(snowballs in hell type chance) that it will get less mosaic when you flash
cool
Hi
One of the easiest things to try is in-situ proteolysis - chymotrypsin (1:1000
or so, we add 1ul of a 1mg/ml solution of chymotrypsin in H20 to about 100ul of
protein solution) is the classic - mix the protease with the protein and then
set up as normal. Of course you can try other protease
Hi,
I used the Axygen sealer for more than a year, and it worked fine. A couple of
caveats -
The sealer is cheaper than many automated sealers, but the seals are pre-cut,
and are more expensive.
If you are sloppy laying the seal over the plate it is relatively easy to not
seal one edge (or t
You could try the matrix seeding method, popularised by Allan D'Arcy et al
(Acta Cryst. (2007). D63, 550-554)
Crush up your needles, and use them to seed into a new round of screening. You
may be able to jump into a new crystal system this way
Janet
-Original Message-
From: CCP4 bulle
We have both a Phoenix and a Mosquito - for one-subwell 96 well plate
both are under 3 minutes from the time you put the first droplet down
until you have to seal the plate. The actual protocols take longer
(particularly on the Phoenix) as you have to wash, transfer reservoir
etc.
We manage to fi
Do you want an imager, or a more sophisticated system that will store plates
and image them according to a schedule?
With imaging, it is important to think about what you want out of the system,
as it is easy to be disappointed with them. The images you get will not be as
good as what you see
I bought a replacement Cartesian Tip (in Australia) on the 8th of
october, 2007 and got charged $660 AUD (of which 60 bucks was the GST
tax) - on the 8th of october, the AUD-USD exchange rate was 0.8956, so
that would be $537 USD, which is quite a long way from both '$700' and
'under $300'.
The t
e aliquoted out into 8 separate puddles. The Cartesian was always a
bitch to get running, so given the other two machines, it sits and
collects dust. Anybody want to start negotiating for a (slightly) used
Cartesian?
Janet
Janet Newman
CSIRO Molecular and Health Technologies
343 Royal Pa
The problem with old solutions is not their age as much as they might be
very hard to reproduce, as you won't exactly know what you have. (for
all the reasons that have been mentioned: evapouration, degradation, pH
shifts, mould growth, contamination from people using the wrong pipette
tip in a di
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