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Re: [ccp4bb]: Protein-DNA complex
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You could also fish the crystals out, wash them, and run them on a
denaturing gel. The addition of urea will help to
dissolve your crystals. Depending on the size of your components, a
4-15% Tris-HCl gel may work well. You can stain both the DNA and the
protein with silver. Compare the bands with your controls, run in an
adjacent lane.
John
------------------------------------------------------
John R. Walker, Ph.D.
Memorial Sloan-Kettering Cancer Center
Box 238, 1275 York Avenue
New York, New York
10021
Tel: (212) 639-6713 FAX: (212) 717-3135
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> Laurence Pearl wrote:
> If crystals are big enough/plentiful you could fish some out, wash
> thoroughly and run them into a non-denaturing gel, and stain it with
> ethidium bromide - this binds dsDNA. You can also stain the same gel
> with Coomassie to visualise the protein. Alternatively try and image the
>
> Elias J. Fernandez wrote:
>
> >*** For details on how to be removed from this list visit the ***
> >*** CCP4 home page http://www.ccp4.ac.uk ***
> >
> >Folks,
> >
> >Is there a clever way to tell if our crystals are complexed protein:DNA
> >versus unbound protein, without having to wait until we have maps? We have a
> >fluorophore attached to the DNA fragment.
> >
> >
> >Elias J. Fernandez, PhD
> >Assistant Professor
> >University of Tennessee
> >Biochemistry, Cellular and Molecular Biology
> >Knoxville, TN 37996-0840
> >Tel. No. 865-974-4090
> >Fax. No. 865-974-6306
> >e-mail: elias.fernandez@utk.edu
> >
> >
> >
> >
> >
>
> --
>
> ---------------------------------------------------------------------------
> Laurence H. Pearl
>
> Section of Structural Biology, Institute of Cancer Research
> Chester Beatty Laboratories, 237 Fulham Road
> London SW3 6JB, UK
>
> Phone +44-207-970 6045 : Secretary +44-207-970 6046
> FAX +44-207-970 6051 : E-Mail l.pearl@icr.ac.uk
> ---------------------------------------------------------------------------
> " Live Simply and do Serious Things .. " - Dorothy Crowfoot Hodgkin
> ---------------------------------------------------------------------------
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