Re: [ccp4bb] Microscope camera

[Darren Hart]

Have you considered a Raspberry Pi with camera, or camera mount with 
your existing lens:


https://www.raspberrypi.com/products/#cameras-and-displays

Photos and livestream will be easy to set up. You could try motioneye to 
give you a livestream and take pictures on demand, or at defined time 
intervals.


https://raspberrytips.com/install-motioneye-on-raspberry-pi/

Darren



On 24/04/2024 23:15, Patrick Loll wrote:

Greetings, hive mind,

We have an old (but still useful) Nikon SMZ stereomicroscope that we use for 
mounting crystals. I’d like to attach a digital camera to the phototube, both 
to capture crystal images for archival purposes, and also to live-stream as a 
teaching tool.

I’d be grateful for any suggestions for an inexpensive option here.

When this camera was new we used it with an SLR that captured images on *film* 
(this is where the students gasp). We’ve since gone through one digital camera 
that probably still works, but the interface and software have become 
obsolescent. Meanwhile, the microscope keeps on truckin’; interesting to 
reflect on the relative lifetimes of analog vs. digital tools…

Thanks in advance for any suggestions,

Pat


---
Patrick J. Loll, Ph. D.  (he, him, his)
Professor of Biochemistry & Molecular Biology
Drexel University College of Medicine
Room 10-102 New College Building
245 N. 15th St., Mailstop 497
Philadelphia, PA  19102  USA

(215) 762-7706
pjl...@gmail.com
pj...@drexel.edu



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1

This message was issued to members ofwww.jiscmail.ac.uk/CCP4BB, a mailing list 
hosted bywww.jiscmail.ac.uk, terms & conditions are available 
athttps://www.jiscmail.ac.uk/policyandsecurity/

--

**

Dr. Darren J. Hart,

CNRS Research Director, Institut de Biologie Structurale (IBS)
Unité Mixte de Recherche UMR5075 (CEA-CNRS-UGA)


Director, Integrated Structural Biology Grenoble (ISBG)
Unité d'Appui et de Recherche UAR3518 (CNRS-UGA-CEA-EMBL)

**

Email: darren.h...@ibs.fr
Tel: +33 4 57 42 85 86

Physical address: IBS/ISBG, 71 avenue des Martyrs, 38000 Grenoble, France

Postal address: IBS/ISBG, 71 avenue des Martyrs, CS 20192, 38042 
Grenoble, Cedex 9, France


**




To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1

This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a mailing list 
hosted by www.jiscmail.ac.uk, terms & conditions are available at 
https://www.jiscmail.ac.uk/policyandsecurity/

Re: [ccp4bb] OFFTOPIC question "Two plasmids in one host cell"

[Darren Hart]

The pACYC vectors (probably your p15ori plasmid) is said to have a copy 
number of ~10 copies per cell while your other plasmid is likely in the 
hundreds - see:


https://blog.addgene.org/plasmid-101-origin-of-replication

The protein expression level from the lower copy number plasmid is 
usually a bit lower, but not always.


For sequencing, we use primers that are specific to the pACYC vector so 
it does not matter if there is a second higher copy plasmid present in 
the miniprep.


To increase the amount of pACYC vector for sequencing, you can always 
amplify the copy number by adding spectinomycin to a growing culture - 
this inhibits protein synthesis, but the low copy plasmids continue to 
replicate to hundreds of copies per cell.


Darren



On 30/12/2020 10:35, Anamika Singh wrote:

Hi All,

I have two constructs having different ori, p15ori and M13 ori, 
different promoters araBAD promoter and LacI, and different antibiotic 
resistance chloramphenicol and Ampicillin respectively.
I am managed to get the transformants and getting the expected result 
after blunt digestion with the EcoRV enzyme. Since both the plasmids 
have the site for EcoRV. But the p15 ori has a low copy number that's 
why I am seeing the very faint band as compared to other plasmid in 
the sample.


So I would like to know is there any way that I can quantify the low 
copy number plasmid.
Because I am not able to sequence it with the specific primers it 
could be due to its low concentration.


Please advise.

Thank you.
--
Dr. Anamika Singh
Post-Doctoral Fellow
Silberman Institute of Life Sciences
Hebrew University of Jerusalem, Israel
No: 054-294-8036



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1 




--

**

Dr. Darren J. Hart,

CNRS Research Director, Institut de Biologie Structurale (IBS)
Unité Mixte de Recherche UMR5075 (CEA-CNRS-UGA)


Director, Integrated Structural Biology Grenoble (ISBG)
Unité Mixte de Service UMS3518 (CNRS-UGA-CEA-EMBL)

**

Email: darren.h...@ibs.fr
Tel: +33 4 57 42 85 86

Physical address: IBS/ISBG, 71 avenue des Martyrs, 38000 Grenoble, France

Postal address: IBS/ISBG, 71 avenue des Martyrs, CS 20192, 38042 
Grenoble, Cedex 9, France


**




To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1

This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a mailing list 
hosted by www.jiscmail.ac.uk, terms & conditions are available at 
https://www.jiscmail.ac.uk/policyandsecurity/

Re: [ccp4bb] CCP4BB vs COVID19

[Darren Hart]

Structure of 2019-nCov RNA polymerase:

https://www.biorxiv.org/content/10.1101/2020.03.16.993386v1.full.pdf+html

/Here we report the cryo-EM structure of 2019-nCoV full-length nsp12 in 
complex with cofactors nsp7 and nsp8 at a resolution of 2.9-Å...A 
comparative analysis to show how remdesivir binds to this polymerase is 
also provided. /


Darren



On 22/03/2020 19:18, Nikolay Dobrev wrote:

Dear all,
I assume most of you are aware of the EMBL-EBI datahub which was set 
up in January to provide essential virus research data to all 
scientists, but in case someone missed it I would like to share the link:

https://www.ebi.ac.uk/ena/pathogens/covid-19

You can find all relevant data, from COVID-19 genome sequencing data 
up to x-ray and cryo-EM structures of relevant proteins.


Stay healthy,
Nikolay

*Nikolay Dobrev *
Scientific Officer, Protein Expression and Purification Core Facility
EMBL Heidelberg, Meyerhofstraße 1, 69117 Heidelberg, Germany
T +49 6221 387 8633 | M +49 173 684 0532
twitter.com/EMBLorg  | 
facebook.com/embl.org  | 
youtube.com/user/emblmedia 
Visit www.embl.org/events for a complete 
list of all EMBL events.







On Sun, Mar 22, 2020 at 17:26, DUMAS Philippe (IGBMC) 
 wrote:


Relevant to the discussion:

* Cell, Vol. 110, 551–561, September 6, 2002, Copyright 2002 by
Cell Press
An RNA Thermosensor Controls Expression of Virulence Genes in
Listeria monocytogenes

* Bacterial RNA thermometers: molecular zippers and switches
Jens Kortmann and Franz Narberhaus
NATURE REVIEWS | MICROBIOLOGY VOLUME 10 | APRIL 2012 | 255

*An RNA Thermometer Activity of the West Nile Virus Genomic
30-Terminal Stem-Loop Element Modulates Viral Replication Eciency
during Host Switching
Viruses 2020, 12, 104; doi:10.3390/v12010104

* Temperature triggers immune evasion by Neisseria meningitidis
Edmund Loh1*, Elisabeth Kugelberg2*, Alexander Tracy1, Qian
Zhang2, Bridget Gollan2, Helen Ewles2, Ronald Chalmers3,
Vladimir Pelicic2 & Christoph M. Tang1,2
Nature (2013)

Philippe Dumas

*De: *"James Holton" mailto:jmhol...@lbl.gov>>
*À: *"CCP4BB" mailto:CCP4BB@JISCMAIL.AC.UK>>
*Envoyé: *Dimanche 22 Mars 2020 16:38:28
*Objet: *Re: [ccp4bb] CCP4BB vs COVID19

Thank you Patrick,

RNA structure is still structural biology, so I think relevant
here.  It seems to me that RNA as a thermometer would be an easy
hypothesis to test? Has anyone measured virulence vs temperature
in cell culture?

The 3D structure of the genome is no doublt important.  I wouldn't
want to try crystallizing the whole thing, but I wonder if this
might be an excellent target for cryoEM?  A challenge for that "we
can classify our way out of anything" philosophy?  And the result
would most certainly be interesting.

-James Holton
MAD Scientist

On 3/21/2020 8:41 AM, Patrick Shaw Stewart wrote:


James, this isn't conventional structural biology, but may be
of interest, and I haven't been able get any mainstream
virologists to think about it.

The protein sequences are obviously of interest, but so are
the RNA sequences at both ends of the Covid genome, which have
conserved secondary structure.  A few years ago a paper came
out suggesting that wild-type influenza has multiple "RNA
thermometers", which may play an important role in the tropism
of influenza.  Similar mechanisms may exist in other
respiratory viruses, including Covid.

My take on this, and the relevant papers, are below.

Good luck to everyone and stay well,

Patrick



https://oldwivesandvirologists.blog/Covid-19-and-the-trade-off-model-of-selection/


My paper in /Medical Hypotheses
/http://douglas.co.uk/f_ftp1/ShawStewart_final_1-s2.pdf

Narberhaus, Franz, Torsten Waldminghaus, and Saheli
Chowdhury. "RNA thermometers." /FEMS microbiology
reviews/ 30.1 (2006): 3-16.

Chursov, Andrey, et al. "Specific temperature-induced
perturbations of secondary mRNA structures are associated
with the cold-adapted temperature-sensitive phenotype of
influenza A virus." /RNA biology/ 9.10 (2012): 1266-1274.

Yang, Dong, and Julian L. Leibowitz. "The structure and
functions of coronavirus genomic 3′ and 5′ ends." /Virus
research/ 206 (2015): 120-133.



On Fri, Mar 20, 2020 at 10:59 PM James Holton
mailto:jmhol...@lbl.gov>> wrote:

You might think that as a structural biologist you won't
be able to do
much about COVID-19 anytime soon, but that is not true. 
 

Re: [ccp4bb] Off-topic: (micro)array image analysis software

[Darren Hart]

Hello,

You could run Image Quant TL in a VM (parallels, vmware or virtual box).

https://bmi.cchmc.org/resources/software/imagequant-tl

For this appliation (arrays), we use an old program called VisualGrid 
that is no longer available and run it an isolated XP VM via virtual box 
(in linux).


Darren


On 21/01/2020 15:48, Bärbel Blaum wrote:


Hello,

clearly off-topic but maybe someone here is experienced in the 
analysis of array scans and can offer some advice? That would be 
beautiful. Here’s the problem: I want to test an ELISA-based HT array 
for phosphorylation profiling of a whole pathway, with almost 200 
antibodies at once. The array company offers free scanning of the 
arrays, i.e. we do the experiment, sent the slides back, and receive 
the scans as raw images in tiff format. Can anyone suggest a suitable 
program to analyse such images of array scans on a Mac? There are six 
replicates per antibody per slide so in theory a good basis for at 
least semi-quantitative analysis - but finding a program to analyse 
these data is a real pain. The scans are obtained with a GenePix 
scanner I am being told but the instrument’s software does not run on 
a Mac. I tried ImageJ, which is open source and runs but needs some 
plugin for arrays that does not work for me (I suspect it worked for a 
previous version of ImageJ). I also tried TIGR Spotfinder, which in 
theory seems the perfect program (plenty of documentation), is also 
open source and Mac compatible - but when I try to compile it several 
files seem missing from the sourceforge package (and I cannot find 
another source).


Does anyone use either ImageJ with the array plugin or the Spotfinder 
and can assure me that these options are still being developed or 
maybe knows for sure they are dead and not worth investing any more 
time? Or, even better, could point me to a program that runs on a Mac 
and is suitable for array analysis (it does not actually have to be 
free as long as it works for users who do not write code). I do have a 
GAL file for the images.


Many thanks for your help and sorry for the spam!

Bärbel

--

Bärbel Blaum, PhD

Inthera Bioscience AG

Einsiedlerstrasse 34

CH-8820 Waedenswil

Switzerland

E-Mail: baerbel.bl...@intherabio.com

Phone: +41 43 477 94 72--




To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1


--

**

Dr. Darren J. Hart,

CNRS Research Director, Institut de Biologie Structurale (IBS)
Unité Mixte de Recherche UMR5075 (CEA-CNRS-UGA)

Director, Integrated Structural Biology Grenoble (ISBG)
Unité Mixte de Service UMS3518 (CNRS-UGA-CEA-EMBL)

**

Email: darren.h...@ibs.fr

Tel: +33 4 57 42 85 86

Physical address: IBS/ISBG, 71 avenue des Martyrs, 38000 Grenoble, France

Postal address: IBS/ISBG, 71 avenue des Martyrs, CS 20192, 38042 
Grenoble, Cedex 9, France


**




To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1

Re: [ccp4bb] Propeptide cleavage for complex assembly

[Darren Hart]

Influenza polymerase heterotrimer expressed in insect cells (MultiBac 
technology) as a fully synthetic polyprotein with subunits separated by 
TEV sites



TevProtease-Tev-PA-Tev-PB1-Tev-PB2-Tev-CFP


Reich et al. Nature 2014; doi:10.1038/nature14009


/Extended Data Figure 1 | Production and characterization of influenza B//
//polymerase heterotrimer. a, Schematic of the self-cleaving polyprotein//
//construct used to express recombinant influenza B heterotrimeric 
polymerase//
//in insect cells. N-terminally it encodes the tobacco etch virus (TEV) 
protease//
//that cleaves C-terminal to the amino-acid sequence ENLYFQ (in italics) 
and//

//releases N-terminally His-tagged PA, PB1, C-terminally Strep-tagged PB2//
//and cyan fluorescent protein (CFP) for facilitated expression 
monitoring.//
//Arrows indicate the N-to-C-terminal direction and the termini of each 
mature//

//protein. The histidine and streptavidin tags are underlined./


Darren




On 02/10/2019 10:18, Paula Salgado wrote:

Dear colleagues

We have been working on a protein that is produced as a pro-peptide, 
cleaved internally and reassembled into a complex. The interacting 
regions are the new C and N-termini at the cleavage site, so no 
rearrangement or loss of part of the protein is involved. The 
resulting interacting region is quite intricate, with a helix bundle 
and beta-sheet composed of members of each partner protein.


I wonder if there are other examples of such processing required for 
folding/assembly of a functional complex? I'm not talking of 
processing involving removal of termini or internal regions like 
signal removal in transported proteins or maturation of preproinsulin, 
for example. I'm interested in examples where a propeptide is cleaved, 
then the two proteins reassemble in a complex via interactions of the 
similarly oriented new termini.


Any suggestions most appreciated.

Thanks a lot!
Paula



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1


--

**

Dr. Darren J. Hart,

CNRS Research Director, Institut de Biologie Structurale (IBS)
Unité Mixte de Recherche UMR5075 (CEA-CNRS-UGA)

Director, Integrated Structural Biology Grenoble (ISBG)
Unité Mixte de Service UMS3518 (CNRS-UGA-CEA-EMBL)

**

Email: darren.h...@ibs.fr

Tel: +33 4 57 42 85 86

Physical address: IBS/ISBG, 71 avenue des Martyrs, 38000 Grenoble, France

Postal address: IBS/ISBG, 71 avenue des Martyrs, CS 20192, 38042 
Grenoble, Cedex 9, France


**




To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1

Re: [ccp4bb] PyMOL now packaged as a snap on Linux

[Darren Hart]

You have to start it via the menu item, not the command line.

Darren

On 03/06/2019 14:31, Pedro Matias wrote:


Well, I can't get it to work in FC30 - I installed FC30 on a virtual 
machine and managed to install both snapd and pymol-oss but pymol does 
not run as a command.


Am I missing something rather obvious?

Pedro

Às 12:46 de 03/06/2019, Matic Kisovec escreveu:

Hi,

thank you very much for this. Work fine on Ubuntu 19.04.

Kind regrds,
Matic


On 17. 05. 19 20:04, Arunabh Athreya wrote:

Thanks for sharing this.

Get Outlook for Android <https://aka.ms/ghei36>


*From:* CCP4 bulletin board  on behalf of 
David Schuller 

*Sent:* Thursday, May 16, 2019 11:41:19 PM
*To:* CCP4BB@JISCMAIL.AC.UK
*Subject:* Re: [ccp4bb] PyMOL now packaged as a snap on Linux
It seems to work on Fedora 30 and on Scientific Linux 7 (which means 
it should also work on RHEL7 and Centos 7)


Thank you.



On 5/16/19 10:47 AM, Darren Hart wrote:

Some more info:

https://snapcraft.io/pymol-oss

It seems to work exactly as expected.

Darren


On 16/05/2019 16:05, Folmer Fredslund wrote:

Hi Darren,

That's brilliant!

I'll give it a spin and see how it works.

Best regards
Folmer


tor. 16. maj 2019 13.02 skrev Darren Hart <mailto:darren.h...@ibs.fr>>:


Since yesterday, PyMOL (open source version v2.3) has been
packaged as a
distro-independent "snap" that can be installed easily on linux
platforms - no more cloning from gitlab and compiling after
installing
the dependencies.

On Ubuntu, install from software centre or:

sudo snap install pymol-oss

Many distros have the snap architecture already installed. If
not, you
just need to install snapd in the regular way first (e.g. on
Debian:
sudo apt install snapd).

Hope this is useful for some folks.

Best regards,

Darren



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1



--




To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1



--
===
All Things Serve the Beam
===
David J. Schuller
modern man in a post-modern world
MacCHESS, Cornell University
schul...@cornell.edu



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1




To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1






To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1


--

Industry and Medicine Applied Crystallography
Macromolecular Crystallography Unit
___
Phones : (351-21) 446-9100 Ext. 1669
  (351-21) 446-9669 (direct)
  Fax   : (351-21) 441-1277 or 443-3644

email :mat...@itqb.unl.pt

http://www.itqb.unl.pt/research/biological-chemistry/industry-and-medicine-applied-crystallography
http://www.itqb.unl.pt/labs/macromolecular-crystallography-unit

Mailing address :
Instituto de Tecnologia Quimica e Biologica António Xavier
Universidade Nova de Lisboa
Av. da República
2780-157 Oeiras
PORTUGAL

ITQB NOVA, a great choice for your PhD
https://youtu.be/de6j-aaTWNQ

Master Programme in Biochemistry for Health
https://youtu.be/UKstDCFjYI8



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1



--

**

Dr. Darren J. Hart,

CNRS Research Director, Institut de Biologie Structurale (IBS)
Unité Mixte de Recherche UMR5075 (CEA-CNRS-UGA)

Director, Integrated Structural Biology Grenoble (ISBG)
Unité Mixte de Service UMS3518 (CNRS-UGA-CEA-EMBL)

**

Email: darren.h...@ibs.fr

Tel: +33 4 57 42 85 86

Physical address: IBS/ISBG, 71 avenue des Martyrs, 38000 Grenoble, France

Postal address: IBS/ISBG, 71 avenue des Martyrs, CS 20192, 38042 

Re: [ccp4bb] PyMOL now packaged as a snap on Linux

[Darren Hart]

Some more info:

https://snapcraft.io/pymol-oss

It seems to work exactly as expected.

Darren


On 16/05/2019 16:05, Folmer Fredslund wrote:

Hi Darren,

That's brilliant!

I'll give it a spin and see how it works.

Best regards
Folmer


tor. 16. maj 2019 13.02 skrev Darren Hart <mailto:darren.h...@ibs.fr>>:


Since yesterday, PyMOL (open source version v2.3) has been
packaged as a
distro-independent "snap" that can be installed easily on linux
platforms - no more cloning from gitlab and compiling after
installing
the dependencies.

On Ubuntu, install from software centre or:

sudo snap install pymol-oss

Many distros have the snap architecture already installed. If not,
you
just need to install snapd in the regular way first (e.g. on Debian:
sudo apt install snapd).

Hope this is useful for some folks.

Best regards,

Darren



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1



--




To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1

[ccp4bb] PyMOL now packaged as a snap on Linux

[Darren Hart]

Since yesterday, PyMOL (open source version v2.3) has been packaged as a 
distro-independent "snap" that can be installed easily on linux 
platforms - no more cloning from gitlab and compiling after installing 
the dependencies.


On Ubuntu, install from software centre or:

sudo snap install pymol-oss

Many distros have the snap architecture already installed. If not, you 
just need to install snapd in the regular way first (e.g. on Debian: 
sudo apt install snapd).


Hope this is useful for some folks.

Best regards,

Darren



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1

[ccp4bb] iNEXT annual user meeting Grenoble, 19-21 March 2018

[Darren Hart]

*Announcement of the iNEXT Annual User Meeting in Grenoble*

The* 3**^rd **Annual Users Meeting of iNEXT* will take place from 
*19*^*th * *to 21*^*st* * March 2018*at the European Photon and Neutron 
(EPN) Science campus in *Grenoble (France)*. The EU-funded structural 
biology project *iNEXT* provides free user access to some of the most 
advanced facilities for structural biology in Europe: X-ray synchrotron 
sources, high-field NMR facilities, Electron Microscopy, and other 
imaging and biophysical characterization facilities.


The meeting is intended to bring together iNEXT users and iNEXT partners 
to exchange on recent scientific advances made in the field of 
structural biology. A large part of the meeting is devoted to scientific 
presentations in various fields of structural biology


Confirmed speakers of this scientific meeting include *Christian 
GRIESINGER, *(MPI for Biophysical Chemistry, Göttingen, Germany), *Maria 
SUNNERHAGEN (*Linköbing University, Sweden),*Chris ULENS (*Laboratory of 
Structural Neurobiology,**Leuven, Belgium), *Ilme SCHLICHTING *(MPI for 
Medical Research, Heidelberg, Germany), and *Jürgen PLITZKO (*MPI for 
Biochemistry, Martinsried, Germany). Additional speakers will be 
selected from the submitted abstracts.



*Registration to the event (including lodging on-site and meals) is free 
of charge.***Additional information on the iNEXT AUM meeting 2018 and 
registration at: http://www.esrf.eu/inext-annual-users-meeting-2018.


Deadline for registration is *February**5*^*th* .

The 3^rd  Annual Users Meeting will be followed by a dedicated“*iNEXT 
meets Industry*” workshop 
from 21^st to 23^rd 
 March at the same place (EPN campus, Grenoble).



See also:

http://www.inext-eu.org/3rd-inext-annual-user-meeting-in-grenoble-registration-is-open/

Re: [ccp4bb] ccp4 website not secure

[Darren Hart]

Set up a certificate via letsencrypt and move to https? It is quick, 
easy and free.


https://letsencrypt.org/

Darren



On 11/10/17 18:34, Edward A. Berry wrote:

Is it because of this? Its been coming for a while now:
https://blog.mozilla.org/security/2015/04/30/deprecating-non-secure-http/

I see no reason why a website, dedicated to providing information
available to everyone, should be required to use https.
The web is too focused on e-commerce, where security is more important.

I hope firefox can be configured to allow insecure http, perhaps with 
a warning.

Otherwise downgrade to previous version (FF 47 connects fine).
Just be sure when visiting ccp4.ac.uk, don't enter personal info
like your tax-payer ID or credit card number. If you need to login
to modify the wiki or your subscription details, use another password for
the bank.

eab

On 10/11/2017 05:15 AM, Markus Heckmann wrote:

If anyone from CCP4 website has noticed that...
Your connection is not secure


The owner of www.ccp4.ac.uk has configured their web site improperly.
To protect your information from being stolen, Firefox has not
connected to this web site.


(https warning message)
https://www.ccp4.ac.uk/ccp4online/





--

**

Dr. Darren J. Hart,

CNRS Research Director, Institut de Biologie Structurale (IBS)
Unité Mixte de Recherche UMR5075 (CEA-CNRS-UGA)


Director, Integrated Structural Biology Grenoble (ISBG)
Unité Mixte de Service UMS3518 (CNRS-CEA-UGA-EMBL)

**

Email: darren.h...@ibs.fr
Tel: +33 4 57 42 85 86

Physical address: IBS/ISBG, 71 avenue des Martyrs, 38000 Grenoble, France

Postal address: IBS/ISBG, 71 avenue des Martyrs, CS 20192, 38042 
Grenoble, Cedex 9, France


**




smime.p7s
Description: S/MIME Cryptographic Signature

Re: [ccp4bb] off-topic: bug busting

[Darren Hart]

Adapted from periplasmic fractionation protocol, but for cytoplasmic 
proteins:


1. Incubate resuspended cells with lysozyme in normal buffer but with
   20% sucrose.
2. Spin down sphaeroplasts (unbroken cells with no cell wall - ultra
   fragile). This also removes periplasmic proteases.
3. Resuspend white pellet in buffer of choice and freeze-thaw. Or
   dilute buffer for osmotic shock. Or sonicate *very* lightly -
   perhaps using an aging sonicator!
4. Gives complete lysis from 2 ml to 2 litre culture volumes very
   reproducibly, and avoids proprietary bug-buster type detergent mixes.


See this paper - it also greatly increases purification (10-fold) of low 
abundance his tag proteins that are otherwise outcompeted by periplasmic 
components (siderophores?)


http://www.nature.com/nmeth/journal/v6/n7/full/nmeth0709-477.html

Darren


On 04/02/14 17:49, Phoebe A. Rice wrote:
Some time ago, there was a nice discussion of cost-effective, wimpy 
protein-friendly ways to break open E. coli.  We're thinking about 
replacing an aging sonicator. If people have a favorite gizmo, could 
they repeat that advice?

thank you,
  Phoebe Rice

++

Phoebe A. Rice
Dept. of Biochemistry  Molecular Biology
The University of Chicago

773 834 1723; pr...@uchicago.edu mailto:pr...@uchicago.edu
http://bmb.bsd.uchicago.edu/Faculty_and_Research/

http://www.rsc.org/shop/books/2008/9780854042722.asp



--

**

Dr. Darren J. Hart,

CNRS Research Director, Unit of Virus Host Cell Interactions (UVHCI)
Unité Mixte Internationale UMI 3265 (CNRS-EMBL-UJF)


Director, Integrated Structural Biology Grenoble (ISBG)
Unité Mixte de Service UMS 3518 (CNRS-CEA-UJF-EMBL)

**

Email: h...@embl.fr
Tel: +33 4 76 20 77 68; Fax: +33 4 76 20 71 99

Postal address: UVHCI/ISBG, 6 rue Jules Horowitz, BP181, 38042 Grenoble, 
Cedex 9, France


For funded access to ESPRIT construct screening via EU FP7 BioStruct-X: 
www.biostruct-x.eu/


**

Re: [ccp4bb] Molecular cloning software for linux

[Darren Hart]

Try
http://gentle.magnusmanske.de/

Resembles VectorNTI, is free, and works on Linux, Windows and Mac 
(although I've only used it on XP).


Seems quite powerful, but not polished like the more commercial solutions.

Darren

On 08/02/13 05:16, Theresa Hsu wrote:

Dear all

Is there any good molecular cloning software for linux? It should read plasmid 
constructs in Genbank format, do assembly of plasmid sequencing results, 
display the sequencing trace and highlight regions where there are mutations 
based on relative QV values, has a list of built-in restriction enzyme 
recognition sequences and also has a database holding all the constructs.

It does not have to be free as long as the price is reasonable for single users.

Thank you.

Theresa



--
**
Dr. Darren J. Hart,
Team Leader, High Throughput Protein Lab
European Molecular Biology Laboratory (EMBL) Grenoble Outstation

Director, Integrated Structural Biology Grenoble (ISBG)
Unité Mixte de Service UMS 3518 (CNRS-CEA-UJF-EMBL)

**
www.embl.fr/research/unit/hart/index.html

For funded access to ESPRIT construct screening via EU FP7 BioStruct-X:
http://www.biostruct-x.eu/

Email: h...@embl.fr
Tel: +33 4 76 20 77 68
Fax: +33 4 76 20 71 99
Postal address: EMBL, 6 rue Jules Horowitz, BP181, 38042 Grenoble, Cedex
9, France
**

[ccp4bb] Research Technician Post Doc positions at EMBL Grenoble

[Darren Hart]

Hello,
Two positions available in my lab at EMBL Grenoble:

   1. Post doc position via the BioStruct-X Joint Research Activity (JRA)
   workpackage Mammalian cell toolbox for structural biology (collab. with
   Oxford University).
   2. Research technician position.

Details pasted below. Further info and online application at
www.embl.org/jobs.

Thanks,
Darren


Research technician, High-Throughput Protein Expression

Location: Grenoble, France
Staff Category: Staff Member
Contract Duration: 1 year
Grading: 4 or 5, depending on experience and qualifications
Closing Date: 31 October 2012
Reference number: GR_00044

Job Description
The European Molecular Biology Laboratory (EMBL) is one of the highest
ranked scientific research organisations in the world. The Headquarters
Laboratory is located in Heidelberg (Germany) and the outstations are in
Grenoble (France), Hamburg (Germany), Hinxton (UK) and Monterotondo (Italy).

A technician position is available in the High Throughput Protein
Technologies laboratory of Darren Hart at EMBL Grenoble. It is funded by
P-CUBE and BioStruct-X, European “Research Infrastructures” projects
established to provide access to unique high-throughput protein production
facilities by visiting international researchers.

The candidate will be responsible for molecular biology and automation
tasks associated with high throughput screening of expression constructs in
bacteria using a new robotic technology, ESPRIT. Projects will be conducted
with both local and international visitors, and the candidate will be
responsible for the planning of the work and visits, together with
experimental supervision. In addition to the project support roles, the
candidate will work on a set of defined research activities involving
vector design, protocol development and crystallisation studies.

Tasks include DNA purification and enzymatic manipulation, PCR, operation
of robotics and other advanced instruments, protein expression and
purification. Additional tasks include the development of new protocols,
testing of new equipment for the facility, keeping accurate records of
consumable use on a project basis, ordering of laboratory consumables and
monitoring associated budgets, meeting with visiting representatives from
suppliers, general maintenance of the robotic laboratory equipment and
arranging repairs as necessary.

Qualifications and Experience
The candidate should hold a Masters degree, or have a good level of
previous, relevant work experience. He/she should have a sound experimental
background in DNA manipulation methods, E. coli protein expression and
protein purification. Experience in high throughput expression, protein
engineering, directed evolution and/or laboratory automation would be
advantageous.The candidate should have experience of working in an English
speaking environment.The ability to work with team members and laboratory
visitors is essential. Fluency in English and good computer skills are
mandatory


Postdoctoral Fellowship in Molecular Biology

Location: Grenoble, France
Staff Category: Postdoctoral Fellow
Contract Duration: up to 18 months
Grading: N/A
Closing Date: 31.October 2012
Reference number: GR_00045

Job Description
The European Molecular Biology Laboratory (EMBL) is one of the highest
ranked scientific research organizations in the world. The Headquarters
Laboratory is located in Heidelberg (Germany) and the outstations are in
Grenoble (France), Hamburg (Germany), Hinxton (UK) and Monterotondo (Italy).

EMBL is seeking to recruit a postdoctoral researcher to join the team of
Darren Hart. The position is funded by the European Framework 7 BioStruct-X
project and is a part of a  collaboration to develop a new expression
technology for structural biology that can be applied to challenging human
proteins. The project will combine random library technologies and high
throughput robotics developed at EMBL (PubMed ID: 20206698, 21515383 
21364980) with parallelised mammalian expression approaches from the
Aricescu lab, University of Oxford (PMID: 17001101). The subjects of the
method development will be medically important human proteins where the aim
is to initiate structural and functional studies.

Qualifications and Experience
The successful applicant should hold a Ph.D. in molecular biology and have
experience of transfecting and handling mammalian cell lines, as well as
being a skilled molecular biologist. He/she should be able to work
independently and take responsibility for his/her own project. Strong
teamwork and communication skills are required as well as reliability,
attention to detail and effective time management. Motivation to work in a
multidisciplinary and international environment is fundamental to this
position. Good communication and presentation skills and fluency in English
are expected.

Application Instructions
Please apply online through www.embl.org/jobs

Please note that appointments on fixed term contracts can be renewed,
depending

[ccp4bb] Workshop on Advanced Protein Engineering Techniques 3rd P-CUBE User Meeting, Heidelberg, 22-24 Oct 2012

[Darren Hart]

Workshop on Advanced Protein Engineering Techniques  3rd P-CUBE User
Meeting  Heidelberg, 22 - 24 October 2012*
*
* Please apply here:  www.p-cube.eu*

Registration deadline: 10 August 2012

*3rd P-CUBE User Meeting (22 Oct 2012)*

Get to know everything about the numerous platforms of P-CUBE and listen to
the platform leaders presenting their infrastructure (Protein Expression in
Bacterial  Eukaryotic Cells, MultiBac, ESPRIT, DARPins, High-throughput
Crystallization, Advanced Microscopy). If you are a former user of one of
the platforms, you might even get selected to speak about your data.  All
the others will have the opportunity to present posters. Please register
online for the P-CUBE User Meeting at www.p-cube.eu.

*Workshop on Advanced Protein Engineering Techniques (23 - 24 Oct 2012)*

The Workshop on Advanced Protein Engineering Techniques is targeted at
early career scientists (PhD student/Postdoc), who are interested in
learning the basics of advanced protein engineering techniques.

 The hands-on course will be taught in small groups and will cover the
following topics:

•Genetically encoding unnatural amino acids via amber
suppression for site-specific protein engineering

•Advanced protein engineering with intein technology

Speakers include:

Stefan Knapp, Oxford University, UK (Keynote Lecture)

Luc Brunsveld, TU Eindhoven, NL
Maja Köhn, EMBL Heidelberg, DE

Edward Lemke, EMBL Heidelberg, DE

Henning Mootz, Muenster University, DE

Daniel Summerer, Konstanz University, DE


To apply for the P-CUBE User Meeting and the workshop please visit our web
page at www.p-cube.eu. Scientists applying for the workshop will be asked
to submit their CV as well as a letter of motivation. A maximum of 12
participants will be selected for the workshop. The meeting  workshop will
be open to scientists from EU Member and/or Associated States.

*General Information*

•There is no registration fee.

•Lunch  2 dinners will be paid by P-CUBE.

•Accommodation will be covered for workshop participants as
well as selected speakers at the user meeting in the Hotel ISG (shared
rooms, 3 nights max. workshop participants) and the Exzellenz Hotel (shared
rooms, 2 nights max. user meeting speakers).

•Participants are responsible for their own travel costs.
However, travel costs of speakers at the user meeting will be partially
reimbursed (300 € max.). For workshop participants there is the possibility
to apply for a small number of travel grants (300 € max.).

*Venue*

The 3rd P-CUBE User Meeting and the workshop will take place at EMBL
Heidelberg, Meyerhofstraße 1, 69117 Heidelberg, Germany. Please visit this
webpage: www.embl.de

*For more information, please contact Petra Lindemann (
petra.lindem...@embl.de) or Jutta Tatzel (j.tat...@bioc.uzh.ch)*

We look forward to welcoming you in Heidelberg!
Markus Grütter (University of Zurich, CH)
Maja Köhn (EMBL Heidelberg, DE)
Edward Lemke (EMBL Heidelberg, DE)




Dr. Petra Lindemann
Scientific Administrator

Mueller (Christoph) Group
EMBL Heidelberg
Meyerhofstraße 1
69117 Heidelberg
Germany
phone: +49 6221 387- 8365

Re: [ccp4bb] Expressed protein hinders cell lysis?

[Darren Hart]

As a suggestion:


   1. Perform your lysozyme treatment in buffer with 20% sucrose.
   2. Then pellet your still intact cells and remove supernatant (cells
   pellet is white and cell lack walls - sphaeroplasts). Periplasmic
   proteases,  metallophores, etc are all washed away.
   3. Then resuspend in your lysis buffer and sonicate - cells are very
   fragile and burst easily.


For more details, see supp methods of this paper:

Nat Methods. 2009 Jul;6(7):477-8.
Enabling IMAC purification of low abundance recombinant proteins from E.
coli lysates.
Magnusdottir A, Johansson I, Dahlgren LG, Nordlund P, Berglund H.
PMID:19564847 [PubMed - indexed for MEDLINE]

Good luck,
Darren




On 21 June 2012 14:44, J. Valencia S. valen...@gene.nagoya-u.ac.jp wrote:

 Greetings, everyone. We need to ask your advice on an issue with one of
 our proteins expressed in E. coli Rosetta cells. This yeast-derived
 protein has a very low yield compared to others we work with, and we think
 it is because the cells are hard to lyse: even after 3 cycles in a cell
 cracker the solution barely changes colour.

 We have no problems lysing Rosetta cells expressing other yeast-derived
 soluble proteins, and we usually obtain enough for our crystallisation
 screens. For the aforementioned protein we have already tried using STAR
 cells, varying the contents of the lysis buffer, sonicating, or adding
 FeSO4 to the solution (we think the protein binds Fe or Mn because it is
 yellow), but to no avail.

 Searching the ccp4bb archive and other resources did not help, so we would
 like to ask 2 questions to the community in order to focus our efforts
 better:
 1. How can a recombinant protein make a cell harder to lyse?
 2. Do you have any suggestions to avoid this effect?

 We appreciate any input, and will be sure to post a summary for future
 reference once this issue is solved.

 Sincerely,


 --
 J. Valencia S.
 PhD student
 CGR-NU




-- 
**
Dr. Darren Hart,
Team Leader
High Throughput Protein Lab
Grenoble Outstation
European Molecular Biology Laboratory (EMBL)
**
www.embl.fr/research/unit/hart/index.html

For funded access to ESPRIT construct screening via EU FP7 PCUBE:
http://tinyurl.com/ydnrwg4

Email: h...@embl.fr
Tel: +33 4 76 20 77 68
Fax: +33 4 76 20 71 99
Skype: hartdarren
Postal address: EMBL, 6 rue Jules Horowitz, BP181, 38042 Grenoble, Cedex
9, France
**

Re: [ccp4bb] Expression of Viral proteins for crystallography

[Darren Hart]

I think the explanation is this:
The source is natural viral RNA which is a mixture of naturally mutated
sequences (e.g. flu forms such a quasispecies)
See:
http://www.virology.ws/2009/05/11/the-quasispecies-concept/

The pooled RNA has an average sequence that you see when you sequence the
pooled cDNA (individual mutations are hidden by the averaging effect of
having many sequences present).

But when you clonally separate DNA molecules by transformation (1 plasmid
enters 1 cell to yield 1 colony), you see each individual molecule
represented 100% in the sequencing chromatogram from the plasmid DNA that
you have isolated from colonies.

This is effect is commonly observed when sequencing influenza virus
isolates from patients. It will have nothing to do with the E. coli strain.
You can avoid it completely by using gene synthesis.

Darren


2012/1/24 Rubén Sánchez Eugenia ruben...@hotmail.com

 Dear everyone,

 I am trying to clone a viral protein in the E. Coli BSJ strain and i am
 having some problems.

 I start from the viral RNA carrying out a reverse transcription and PCR
 (RT-PCR) to obtain the protein cDNA. When I sequence this cDNA to check for
 mutations, there are no mutations. So the RT-PCR works fine.

 Then, I digest the cDNA and I ligate it with a pET plasmid to transform
 the E. Coli BSJ strain. I get recombinant colonies (checked by colony-PCR)
 but when I sequence them I get various mutations (aprox. 2 miss-sense) on
 the inserted cDNA. Furthermore, these mutations are different among
 different transformations and even among colonies of the same plate (in the
 same transformation).

 Maybe these mutations are produced by the cell (because of the lack of
 mutations in the cDNA) but these E. Coli clonning strains are supposed to
 be optimized to prevent the insertion of mutations. So I have no idea
 about what may be the problem.

 I hope you could help me. Thank you.

 Best regards,

 --
 ---
 Rubén Sánchez





-- 
**
Dr. Darren Hart,
Team Leader
High Throughput Protein Lab
Grenoble Outstation
European Molecular Biology Laboratory (EMBL)
**
www.embl.fr/research/unit/hart/index.html

For funded access to ESPRIT construct screening via EU FP7 PCUBE:
http://tinyurl.com/ydnrwg4

Email: h...@embl.fr
Tel: +33 4 76 20 77 68
Fax: +33 4 76 20 71 99
Skype: hartdarren
Postal address: EMBL, 6 rue Jules Horowitz, BP181, 38042 Grenoble, Cedex
9, France
**

Re: [ccp4bb] Off topic: vector map editing and DNA sequence alignment software

[Darren Hart]

://gentle.magnusmanske.de/ it is
pretty similar to Vector NTI (and open source for the ambitious).

For Macs:

Jovine Luca - *DNA Strider *(1.4) runs just fine on both Tiger and Leopard.
For more info, you can contact the author directly: christian.marck_at_cea.
Fr

You could also try Christian Biertuempfel's suggestion of *pDRAW32*: if it
works under Wine on Linux, it should work under Wine (or the commercial
equivalent Codeweavers Crossover) on OS X as well.

Roger Dodd - *PlasmaDNA *which seems pretty good for the basics
http://research.med.helsinki.fi/plasmadna/ . It works under Mac OS X and
Windows... and probably Wine on Linux too.

Jann Sterxx - *Geneouis *also runs on OS X. From my experience (past two
years), the program works just fine. You can download the trial at
http://www.apple.com/downloads/macosx/math_science/index1.html (second
page).





On 27 September 2011 22:36, Sampson, Jared jared.samp...@nyumc.org wrote:

 Hi Florian -

 I've used Geneious for a few years now and been pleased with it.  Also a
 freemium business model: Basic version is free, and Pro version price
 depends on the term and type of license (student, academic/government, or
 commercial).  I find the Basic version suits my limited molecular biology
 needs pretty well. They also have occasional Geneious Days (today happens
 to be one!) when the Basic version can use all the features of the Pro
 version.

 Available for Linux/Mac/Windows in both 32- and 64-bit.

 http://www.geneious.com/

 --
 Jared Sampson
 Xiangpeng Kong Lab
 NYU Langone Medical Center
 550 First Ave MSB 329
 New York, NY 10016
 212-263-7898

 On Sep 27, 2011, at 2:32 PM, Luca Jovine wrote:

 Hi Florian,

 Have a look at Serial Cloner - it's free, runs on OS X, Linux and Windows,
 and is really quite powerful - including the ability to export single or
 multiple sequences to FASTA format text files (however, it can only align
 two sequences at the time I'm afraid):

 http://serialbasics.free.fr/Serial_Cloner.html

 HTH, Luca

 
 Luca Jovine, Ph.D.
 Assistant Professor  EMBO Young Investigator
 Karolinska Institutet
 Department of Biosciences and Nutrition  Center for Biosciences
 Hälsovägen 7, SE-141 83 Huddinge, Sweden
 Voice: +46.(0)8.524-81136  FAX: +46.(0)8.6081-501
 E-mail: luca.jov...@ki.se
 W3: http://jovinelab.org
 

 On 27 Sep 2011, at 17:42 , Florian Schmitzberger wrote:

 Dear All,

 What type of software are people commonly using these days for
 vector/plasmid map editing, making/visualizing vector maps, and aligning
 (small to medium size) DNA sequencing data? Preferably, it should not be too
 expensive and be able to write text files, readable by other programs.

 I am familiar with VectorNTI, which is great for vector visualization and
 editing; but I find it somewhat expensive. Sequencher seems good to quickly
 align DNA sequences (such as from DNA sequencing) with templates, but is not
 free. I have been using ApE for while for alignments, but aligning many
 sequences is more cumbersome than in Sequencher; I have not tested if
 Sequencher is good at visualizing and editing plasmid maps.

 Ideally, I would like to have a single program for both purposes (vector
 editing and DNA sequence comparison). Does something like that exist? What
 are the alternatives to above programs?

 Thank you in advance.

 Florian

   ---
 Florian Schmitzberger, PhD
 Biological Chemistry and Molecular Pharmacology
 Harvard Medical School
 250 Longwood Avenue, Seeley G. Mudd 123
 Boston, MA 02115, US
 Tel: 001 617 432 5603















  
 This email message, including any attachments, is for the sole use of the
 intended recipient(s) and may contain information that is proprietary,
 confidential, and exempt from disclosure under applicable law. Any
 unauthorized review, use, disclosure, or distribution is prohibited. If you
 have received this email in error please notify the sender by return email
 and delete the original message. Please note, the recipient should check
 this email and any attachments for the presence of viruses. The organization
 accepts no liability for any damage caused by any virus transmitted by this
 email.
 =




-- 
**
Dr. Darren Hart,
Team Leader
High Throughput Protein Lab
Grenoble Outstation
European Molecular Biology Laboratory (EMBL)
**
www.embl.fr/research/unit/hart/index.html

For funded access to ESPRIT construct screening via EU FP7 PCUBE:
http://tinyurl.com/ydnrwg4

Email: h...@embl.fr
Tel: +33 4 76 20 77 68
Fax: +33 4 76 20 71 99
Skype: hartdarren
Postal address: EMBL, 6 rue Jules Horowitz, BP181, 38042 Grenoble, Cedex
9

Re: [ccp4bb] Zotero style

[Darren Hart]

You can use this

http://www.somwhere.org/csl/

to build your style.

Darren



On 4 May 2011 09:05, Robbie Joosten robbie_joos...@hotmail.com wrote:

  Hi Everyone,

 Does anyone have a Zotery style template for Acta Cryst and the like, (s)he
 wishes to share? I cannot find it in the repository, but perhaps someone has
 made one for private use.

 Cheers,
 Robbie Joosten

 Biochemistry
 Netherlands Cancer Institute




-- 
**
Dr. Darren Hart,
Team Leader
High Throughput Protein Lab
Grenoble Outstation
European Molecular Biology Laboratory (EMBL)
**
www.embl.fr/research/unit/hart/index.html

For funded access to ESPRIT construct screening via EU FP7 PCUBE:
http://tinyurl.com/ydnrwg4

Email: h...@embl.fr
Tel: +33 4 76 20 77 68
Fax: +33 4 76 20 71 99
Skype: hartdarren
Postal address: EMBL, 6 rue Jules Horowitz, BP181, 38042 Grenoble, Cedex
9, France
**

[ccp4bb] Summary: Web or e-tools for booking instrument time

[Darren Hart]

 mistakenly  remove all
entries from the server if the sync is not done properly. To overcome this
potential hazard, we routinely backup from time to time the current Gcal of
every device.
Hoped I helped!
Chen

Also:
www.prog4biz.com
The application we have developing for managing core facilities, Bookit, has
been evolving tremendously in the past 2 years and many modules and features
are included now, allowing 3 universities to benefit from its use.

I’ve used schedule it software which is available as open source.

http://www.php.brickhost.com/



hope it’s of use,



best regards,

Paul.



-- 
**
Dr. Darren Hart,
Team Leader
High Throughput Protein Lab
Grenoble Outstation
European Molecular Biology Laboratory (EMBL)
**
www.embl.fr/research/unit/hart/index.html

For funded access to ESPRIT construct screening via EU FP7 PCUBE:
http://tinyurl.com/ydnrwg4

Email: h...@embl.fr
Tel: +33 4 76 20 77 68
Fax: +33 4 76 20 71 99
Skype: hartdarren
Postal address: EMBL, 6 rue Jules Horowitz, BP181, 38042 Grenoble, Cedex
9, France
**

[ccp4bb] Off Topic: Web or e-tools for booking instrument time

[Darren Hart]

Hello,
Since we are now part of the Facebook Generation (or at least 10% of the
planet is), it seems there must be a better way of distributing time on our
various Aktas to an institute of scientists than our current system which
involves a race for the paper booking sheet every Thursday morning.
Inevitably people book in a speculative manner and time slots are not always
used due to problems in preparative steps etc. Bits of paper in different
buildings make it difficult to see who booked what and when.

Has anyone successfully implemented an electronic system, perhaps shared
through the cloud, that allows real time booking in a manner that can be
revised easily and has worked well? An electronic calendar (Google..) would
partially address this, but it seems some sort of social networking
site/tool might be better for negotiating instrument time, swapping
timeslots, reallocating unused time if the sample prep fails.

Any suggestions would be greatly appreciated. If governments can be toppled
through Facebook, I'm sure we can book our Aktas in a better way!

Darren

-- 
**
Dr. Darren Hart,
Team Leader
High Throughput Protein Lab
Grenoble Outstation
European Molecular Biology Laboratory (EMBL)
**
www.embl.fr/research/unit/hart/index.html

For funded access to ESPRIT construct screening via EU FP7 PCUBE:
http://tinyurl.com/ydnrwg4

Email: h...@embl.fr
Tel: +33 4 76 20 77 68
Fax: +33 4 76 20 71 99
Skype: hartdarren
Postal address: EMBL, 6 rue Jules Horowitz, BP181, 38042 Grenoble, Cedex
9, France
**

Re: [ccp4bb] software for predicting protein solubility, stabililty and disorders

[Darren Hart]

Run your sequence through these predictors to get an idea of domain location
and disorder.

Disorder predictor links
http://dis.embl.de/
http://globplot.embl.de/
http://www.strubi.ox.ac.uk/RONN
http://bip.weizmann.ac.il/fldbin/findex
http://www.ist.temple.edu/disprot/predictorVSL2.php
http://iupred.enzim.hu/
http://www.pondr.com/
http://bioinf.cs.ucl.ac.uk/disopred/disopred.html

Darren

On 29 July 2010 11:24, Albert Guskov albert.gus...@googlemail.com wrote:

 Hi Vikrant,
 I guess  Xtalpred server might be of interest for you.
 check it at http://ffas.burnham.org/XtalPred-cgi/xtal.pl
 Cheers,
 *Albert GUSKOV (Dr) *| Research Fellow | Division of Structural 
 Computational Biology | Nanyang Technological University
 Proteos 7-01, Biopolis Drive 61, Singapore 138673 Tel: (65) 6586-9690
 GMT+8h | Cell: (65) 8366-2779 | Email: a.gus...@ntu.edu.sg | Web:
 www.ntu.edu.sg

 2010/7/29 vikrant saa powervikr...@yahoo.co.in

  I want to do cloning of  a 40 Kd protein in pRSETA, and pGEX-KT vector.
 I don't have any idea about protein solubility, its multimeric
 form, stability  and disorder etc. There is nothing known in the literature
 also. Is there any software that can predict  these parameters, so that i
 can decide which domain i need to  clone for  soluble and stable protein
 purification.
 *Vikrant
 ***

 
 ***Junior Research Fellow (CSIR) *
 *Lab No. 101, Dr. Varma Lab*
 *Cancer Research Institute
 Advance Centre for Treatment, Research and Eduction in Cancer (ACTREC)
 Tata Memorial Hospital, Kharghar, Navi Mumbai-410210 *
 #







-- 
**
Dr. Darren Hart,
Team Leader
High Throughput Protein Lab
Grenoble Outstation
European Molecular Biology Laboratory (EMBL)
**
www.embl.fr/research/unit/hart/index.html

For funded access to ESPRIT construct screening via EU FP7 PCUBE:
http://tinyurl.com/ydnrwg4

Email: h...@embl.fr
Tel: +33 4 76 20 77 68
Fax: +33 4 76 20 71 99
Skype: hartdarren
Postal address: EMBL, 6 rue Jules Horowitz, BP181, 38042 Grenoble, Cedex
9, France
**

[ccp4bb] OT: Crystallisation compatible detergents

[Darren Hart]

Hello,
We can make a nice 1:1 complex between two proteins that then gradually
precipitates during storage at 4 degC. With 0.005% Tween-20 the complex is
stable and does not preciptate noticeably. We have used this in buffers to
measure the kinetics of binding by Biacore so it is clearly compatible with
functionality.

Is Tween-20 at this concentration compatible with crystallisation? Is it
worth giving a go, or a waste of time? Should we try and remove it, or
reprepare the complex in an alternative detergent?

If we need to try an alternative, what would people recommend?

Thanks in advance for your suggestions,
Darren

-- 
**
Dr. Darren Hart,
Team Leader
High Throughput Protein Lab
Grenoble Outstation
European Molecular Biology Laboratory (EMBL)
**
EMBL webpages:
http://tinyurl.com/6xdltl
http://tinyurl.com/667jrp

Email: h...@embl.fr
Tel: +33 4 76 20 77 68
Fax: +33 4 76 20 71 99
Skype: hartdarren
Postal address: EMBL, 6 rue Jules Horowitz, BP181, 38042 Grenoble, Cedex
9, France
**

[ccp4bb] OT: VectorNTI alternatives - SUMMARY

[Darren Hart]

: christian.marck_at_cea.
Fr

You could also try Christian Biertuempfel's suggestion of *pDRAW32*: if it
works under Wine on Linux, it should work under Wine (or the commercial
equivalent Codeweavers Crossover) on OS X as well.

Roger Dodd - *PlasmaDNA *which seems pretty good for the basics
http://research.med.helsinki.fi/plasmadna/ . It works under Mac OS X and
Windows... and probably Wine on Linux too.

Jann Sterxx - *Geneouis *also runs on OS X. From my experience (past two
years), the program works just fine. You can download the trial at
http://www.apple.com/downloads/macosx/math_science/index1.html (second
page).


-- 
**
Dr. Darren Hart,
Team Leader
High Throughput Protein Lab
Grenoble Outstation
European Molecular Biology Laboratory (EMBL)
**
EMBL webpages:
http://tinyurl.com/6xdltl
http://tinyurl.com/667jrp

Email: h...@embl.fr
Tel: +33 4 76 20 77 68
Fax: +33 4 76 20 71 99
Skype: hartdarren
Postal address: EMBL, 6 rue Jules Horowitz, BP181, 38042 Grenoble, Cedex
9, France
**

Re: [ccp4bb] OT: VectorNTI alternatives

[Darren Hart]

Hello,
I spoke to Invitrogen (France) today and they said that, if asked, they
would provide a 2 month free license so that people could do exactly this
and recover their files.

The also said they would send me details on a simple procedure to extract
the data from a locked version which I will post on the bb.

They told me they have 100,000 users - so hardly a minor specialist
application! At €650-€4000 per license, that looks quite fruitful for them.

An earlier poster made the point that it is a good thing to pay for good
software. I agree and don't want to seem like a moaner, but I object to the
strategy employed here on this occasion. Hence my original post to find out
what the alternatives are.

Darren

2009/1/30 Jeffrey Wilson wil...@ucmail.uc.edu

 On Jan 28, 2009, at 3:47 AM, Darren Hart wrote:


 ps anyone using VNTI might consider a backup of their work by exporting
 files to .gb format. I don't know if a locked up (expired) version permits
 this and you will have no notice that it is about to expire.


 My license recently expired.  I had been running VectorNTI on my Mac
 through a Windows virtual machine.  When the license ran out, I was unable
 to export any of the sequences that I had created.  Now, if there was only a
 way to turn back time so that my computer thought it was still 2008 ... ehm
 ...  ;)

 Jeff




-- 
**
Dr. Darren Hart,
Team Leader
High Throughput Protein Lab
Grenoble Outstation
European Molecular Biology Laboratory (EMBL)
**
EMBL webpages:
http://tinyurl.com/6xdltl
http://tinyurl.com/667jrp

Email: h...@embl.fr
Tel: +33 4 76 20 77 68
Fax: +33 4 76 20 71 99
Skype: hartdarren
Postal address: EMBL, 6 rue Jules Horowitz, BP181, 38042 Grenoble, Cedex
9, France
**

[ccp4bb] OT: VectorNTI alternatives

[Darren Hart]

Hello,
After several years of offering the molecular biology software VectorNTI
free to the academic community (their open access program) and building
up a huge user base, Invitrogen have suddenly announced that they will no
longer renew these free licences and the existing ones will be left to
expire within the year. There are heavy renewal fees for anyone wishing to
continue use of this software.

Can anyone recommend decent alternative PC compatible alternatives? Main
uses are construct and primer design, plus simple quick alignments,
sequence data analysis etc. The database structure for storing sequences
was pretty useful also.

Ideally free, otherwise reasonably priced. I've seen CLCbio and Geneious
have products, both free and paid. Any experience?

Thanks,
Darren
EMBL Grenoble

ps anyone using VNTI might consider a backup of their work by exporting
files to .gb format. I don't know if a locked up (expired) version permits
this and you will have no notice that it is about to expire.

Re: [ccp4bb] OT: VectorNTI alternatives

[Darren Hart]

Hello,
Thanks for all the emails (only some of which reached the bb). It is clear
that I am not alone in feeling aggrieved by Invitrogen suddenly introducing
licence fees, having first persuaded us to put our files and time into their
free product over several years.

Once the thread goes quiet, I'll summarise the suggestions for everyone's
benefit. I've tried several packages today but it would be good if
suggestions were accompanied by brief strengths and weaknesses.

From a first view, there are a number of clunky programs that do the job.
And some that are really just viewers that are difficult to use for sequence
manipulation. As a lab with hundreds of constructs and primers in our
database, we also appreciate the file arrangement/storage as well as the
sequence analysis function.

Thanks
Darren


2009/1/28 Darren Hart h...@embl.fr

 Hello,
 After several years of offering the molecular biology software VectorNTI
 free to the academic community (their open access program) and building
 up a huge user base, Invitrogen have suddenly announced that they will no
 longer renew these free licences and the existing ones will be left to
 expire within the year. There are heavy renewal fees for anyone wishing to
 continue use of this software.

 Can anyone recommend decent alternative PC compatible alternatives? Main
 uses are construct and primer design, plus simple quick alignments,
 sequence data analysis etc. The database structure for storing sequences
 was pretty useful also.

 Ideally free, otherwise reasonably priced. I've seen CLCbio and Geneious
 have products, both free and paid. Any experience?

 Thanks,
 Darren
 EMBL Grenoble

 ps anyone using VNTI might consider a backup of their work by exporting
 files to .gb format. I don't know if a locked up (expired) version permits
 this and you will have no notice that it is about to expire.





-- 
**
Dr. Darren Hart,
Team Leader
High Throughput Protein Lab
Grenoble Outstation
European Molecular Biology Laboratory (EMBL)
**
EMBL webpages:
http://tinyurl.com/6xdltl
http://tinyurl.com/667jrp

Email: h...@embl.fr
Tel: +33 4 76 20 77 68
Fax: +33 4 76 20 71 99
Skype: hartdarren
Postal address: EMBL, 6 rue Jules Horowitz, BP181, 38042 Grenoble, Cedex
9, France
**

[ccp4bb] Workshop: Directed Evolution Approaches in Structural Biology

[Darren Hart]

Directed Evolution Approaches in Structural Biology
Wed 30th  Thu 31st January 2008 at EMBL Grenoble, France.
Funded by: SPINE2COMPLEXES  TEACH-SG (EU FP6)

Info  Register: www-db.embl.de/jss/EmblGroupsOrg/conf_91
See also: www.spine2.eu/SPINE2/meetings/index.jsp?m=28

A number of laboratories are now developing or using library-based 
strategies in combination with structural biology. This may be to 
overcome problems in the structure solution process (expression, 
crystallisation etc.) or to study/engineer proteins for structural and 
functional studies. We aim to strengthen links between these labs and 
provide training for scientists interested in this field.


Topics
1.  Library strategies for “difficult-to-express” proteins
2.  Scaffold proteins and binders as cocrystallisation chaperones
3.  Combinatorial approaches to protein complex definition and expression
4.  Structural studies of in vitro evolved proteins

·Colony filtration (CoFi).
·Combinatorial Domain Hunting (CDH).
·Efficient fragmentation, truncation  mutation approaches
·Robotics  truncation libraries.
·GFP solubility screening.
·Screening by FACS and fluorescent colony picking.
·DARPins  ribosome display.
·DNA shuffling/Oil-in-water emulsions.
·Use of display technologies

Speakers
Stephanie Cabantous Waldo Lab, Los Alamos, USA
Thomas HuberPlückthun Lab, University of Zurich
Speaker Nordlund Lab, Karolinska Institute, Sweden
Renos Savva Domainex Ltd, UK
Darren Hart EMBL Grenoble, France
Martin WalshMRC France, Grenoble
Amir AharoniWeizmann Institute, Israel
Sabine Mazaleyrat   AstraZeneca (ex), UK
Chris Meier UCB-Celltech, UK




--
**
Dr. Darren Hart,
Team Leader
High Throughput Group
Grenoble Outstation
European Molecular Biology Laboratory (EMBL)
**

http://www.embl.fr/groups/htt/expression.html

Email: [EMAIL PROTECTED]
Tel: (33) 4 76 20 77 68
Fax: (33) 4 76 20 71 99
Postal address: EMBL, 6 rue Jules Horowitz, BP181, 38042 Grenoble, Cedex
9, France
**