Re: [ccp4bb] Can nucleoprotein complex inhibit EtBr binding?

2015-02-13 Thread Sylvia Fanucchi
What is the charge on your protein? If it is a very positively charged protein 
it may not enter the gel. I have found this before in EMSAs but normally at low 
protein concentrations it still enters the gel only at the higher concs it does 
not. I had to change protein to DNA ratio and also the percentage of the gel. 
There was quite a bit of optimization that had to be done.

Sent from my iPhone

> On 13 Feb 2015, at 11:50 PM, "Sudipta Bhattacharyya" 
>  wrote:
>
> Dear Community,
>
> Sorry for this off topic question. I am dealing with a non specific DNA 
> binding protein. In a non radio-labelled EMSA DNA:protein titration 
> experiment when I EtBr stained the 5% native acrylamide gel, I could see the 
> DNA control (101 bps) but as the amount of my protein increases I saw the 
> free DNA band getting thinner and thinner but I could not see any band shift 
> and probably all the complex stuck at wells. But, even if I assume the whole 
> DNA length  is occupied by my protein (I determined the ratio to be 1:15 ish) 
> the molecular weight should not reach the molecular weight of the highest DNA 
> marker band (20kbps = 13,200 kDa). Then why I could not see a retarded band 
> in the gel? Or is it forming a EtBr resistant nucleoprotein filament? Could 
> you guys please give me an idea/suggestions?
>
> Many thanks in advance...!!!
>
> My best regards,
> Sudipta.


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Re: [ccp4bb] Can nucleoprotein complex inhibit EtBr binding?

2015-02-13 Thread Pius Padayatti
Sudipta,
the link for the cyanin dyes are at the following link
http://www.lifetechnologies.com/us/en/home/references/molecular-probes-the-handbook/nucleic-acid-detection-and-genomics-technology/nucleic-acid-stains.html

Sorry for the wrong link
P

On Fri, Feb 13, 2015 at 1:50 PM, Sudipta Bhattacharyya <
sudiptabhattacharyya.iit...@gmail.com> wrote:

> Dear Community,
>
> Sorry for this off topic question. I am dealing with a non specific DNA
> binding protein. In a non radio-labelled EMSA DNA:protein titration
> experiment when I EtBr stained the 5% native acrylamide gel, I could see
> the DNA control (101 bps) but as the amount of my protein increases I saw
> the free DNA band getting thinner and thinner but I could not see any band
> shift and probably all the complex stuck at wells. But, even if I assume
> the whole DNA length  is occupied by my protein (I determined the ratio to
> be 1:15 ish) the molecular weight should not reach the molecular weight of
> the highest DNA marker band (20kbps = 13,200 kDa). Then why I could not see
> a retarded band in the gel? Or is it forming a EtBr resistant nucleoprotein
> filament? Could you guys please give me an idea/suggestions?
>
> Many thanks in advance...!!!
>
> My best regards,
> Sudipta.
>



-- 
P


[ccp4bb] Can nucleoprotein complex inhibit EtBr binding?

2015-02-13 Thread Sudipta Bhattacharyya
Dear Community,

Sorry for this off topic question. I am dealing with a non specific DNA
binding protein. In a non radio-labelled EMSA DNA:protein titration
experiment when I EtBr stained the 5% native acrylamide gel, I could see
the DNA control (101 bps) but as the amount of my protein increases I saw
the free DNA band getting thinner and thinner but I could not see any band
shift and probably all the complex stuck at wells. But, even if I assume
the whole DNA length  is occupied by my protein (I determined the ratio to
be 1:15 ish) the molecular weight should not reach the molecular weight of
the highest DNA marker band (20kbps = 13,200 kDa). Then why I could not see
a retarded band in the gel? Or is it forming a EtBr resistant nucleoprotein
filament? Could you guys please give me an idea/suggestions?

Many thanks in advance...!!!

My best regards,
Sudipta.


[ccp4bb] CCP4-6.5 Update 003

2015-02-13 Thread charles . ballard
Dear CCP4 Users

An update for the CCP4-6.5 series has just been released, consisting
of the following changes

 * refmac (all)
 - bug fix for occupancy refinement

 * imosflm/mosflm
 - update to 7.1.2
 - bug fix update

 * pointless (all platforms)
 - update to 1.9.25
 - Some fixes for Saint files. Fix to long-standing but rare bug in hash table
 - Improvements to analysis of 6-fold screws (though still needs further work), 
including avoiding an occasional crash

 * aimless (all)
 - update to 0.5.2
 - bug fix for already merged data

 * feckless (all)
 - update to 0.0.5
 - correction to hash

 * ctruncate (all)
 - update to 1.16.9
 - fix for sigma=0

 * molrep (all)
 - improved multi-copy search

 * sfcheck (all)
 - bug fixes

 * monomer library (all)
 - added missing components

 * ccp4i (all)
 - mrSHELXE renamed to shelxe4mr

 * ccp4.setup (linux, osx)
 - fixed setup scripts for MANPATH issues



Please report any bugs to c...@stfc.ac.uk.

Many thanks for using CCP4.

The CCP4 Core Team

Re: [ccp4bb] OT: PyMOL default opening file type

2015-02-13 Thread Ioan Vancea
Hi,

> On 13 Feb 2015, at 12:31, Yu Wai Chen  wrote:
> 
> Dear all,
> 
> Sorry for this off topic question. When I started PyMOL (Windows 1.6.0.0), it 
> is defaulted to file type ACNT maps. Is there any way that I can set the 
> program to default to PDB? Thanks.

Check this link:

http://www.thewindowsclub.com/change-file-associations-windows 


Cheers,
Ioan



[ccp4bb] OT: PyMOL default opening file type

2015-02-13 Thread Yu Wai Chen

Dear all,

Sorry for this off topic question. When I started PyMOL (Windows 
1.6.0.0), it is defaulted to file type ACNT maps. Is there any way that 
I can set the program to default to PDB? Thanks.


Wai

--
Yu Wai Chen, PhDLecturer
King's College London, Randall Division +44-207-848-8206
New Hunt's House, Guy's Campus, London SE1 1UL, U.K.


Re: [ccp4bb] On the rotamer of Arg

2015-02-13 Thread Eleanor Dodson
If you want to test this, set the ARG atom occupancies to 0.0 - calculate
another map, and see what the density looks like - you can flip through the
aRG rotamers using coot.

Eleanor

On 13 February 2015 at 06:12, Dialing Pretty <
03f1d08ed29c-dmarc-requ...@jiscmail.ac.uk> wrote:

> Dear All,
>
> I have a set of data, in which there are  very nice electron density for
> several Arg residues. However MolProbity analysis demonstrated that the
> percentage of the Arg rotamer in my data is 0%. Although there is the
> possibility that the Arg rotamer in my data is 0%, I still wonder is any
> possibility that to correct the Arg rotamer in my data so that it would
> belong to the scope of commonly observed Arg rotamer.
>
> I am looking forward to getting a reply from you on it.
>
>
> Dialing
>
>
>


[ccp4bb] Macromolecular Crystallography School in Madrid

2015-02-13 Thread Juan A. Hermoso
Dear colleagues,
We are pleased to announce the 

Sixth edition of the Macromolecular Crystallography School 2015, to be held at 
the Institute Química-Física Rocasolano- CSIC (Madrid, Spain).

Dates: May 18 - 23, 2015

Invited speakers and tutors: 

Paul Adams, Berkeley, USA
Pavel Afonine, Berkeley, USA
Armando Albert, Madrid, Spain
Jose Maria Carazo, Madrid, Spain
Kay Diederichs, Konstanz, Germany
Paul Emsley, Cambridge, UK
Carmelo Giacovazzo, Bari, Italy
Xavier Gomis-Ruth, Barcelona, Spain
Juan Hermoso, Madrid, Spain
Eugene Krissinel, Oxford, UK
Rob Nicholls, Cambridge, UK
Maria Solá, Barcelona, Spain
Isabel Usón, Barcelona, Spain

Please find the program of the Workshop, the application form and further 
information, at http://www.xtal.iqfr.csic.es/MCS2015/ 

Applicants:
The workshop has been designed for postgraduate, postdoctoral and research 
scientists that deal with macromolecular crystallography but need further 
knowledge on the routine uses and fundamentals that underlie the modern 
techniques for macromolecular crystallography. The topics will cover 
crystallographic computing and fundamentals using XDS, CCP4 and Phenix, as well 
as the state of the art on topics such as the strategies for protein 
production, phasing with ARCCIMBOLDO, SHELX and SIR2014, modeling with COOT and 
low resolution techniques. MCS2015 will provide five days and a half of 
lectures and workshops. The course is designed for 25 highly motivated students.

Requests to participate must be sent by a free-formated E-mail, before March 
21, 2015 to the following address: cur...@fgua.es, indicating the motivation to 
participate and including a letter of recommendation from PhD Director (only 
for PhD students).
Accepted participants will be notified by e-mail before April 4, 2015. 
Organizers:
Armando Albert (xalb...@iqfr.csic.es ), Juan Hermoso (xj...@iqfr.csic.es) and 
Xavier Gomis-Rüth (xgr...@ibmb.csic.es )

Prof. Dr. Juan A. Hermoso
Dept. Crystallography & Structural Biology
Inst. Quimica-Fisica "Rocasolano"
CSIC (Spanish National Research Council)
Serrano 119, 28006-Madrid, Spain
Phone: +34 917459538
Fax: +34 915642431
xj...@iqfr.csic.es
http://www.xtal.iqfr.csic.es/grupo/xjuan/