In Article <483cft$57r at mserv1.dl.ac.uk>, Andrei Popov
<ANDREI.POPOV at bbsrc.ac.uk> wrote:
>>Does anyone know if in B-cell differentition, the D-J recombination and
>>the Variable Heavy-DJ recombination reactions have been successfully
>>performed in-vitro. A friend of mine said that he thought Dr. Stuart
>>Kauffman of the Santa Fe Institute has done this, but I haven't been
>>able to find any references on this. Any help would be greatly
>>appreciated.
>>No. It has not been done.
>M.Sadofsky (spell?) and coauthers published
>a couple of months ago in Cell a paper on V(D)J recombination
>in vitro but.... it was very far away from total success.
>>
Well, actually I think that it a little closer than no.
M Gehlert's (?) group did have a paper on it, and I think that
it was pretty good, using cell extracts. He presented new
data during the 9th internation congress on immunology showing
that placing recombinant rag I and rag II in a hepes/mops buffer
with a radiolabled RSS 50mer oligo gave a cut product. This isnt
exactly recombination, but shows that the rag s are directly
involved in recombination, and bona fide invitro rec shouldnt be
far off. also, the test for activity of the rag s has been using a
plasmid with rss's in 3t3s with transfected rag i and ii.
this seems to work pretty well. and i wouldnt really call it
invivo.
Regards, Ralph
Ralph M. Bernstein
Dept of Micro/Immuno
University of Arizona
Ph: 602 626 2585
Fx: 602 626 2100
url: http://lamprey.medmicro.arizona.eduhttp://radon.gas.uug.arizona.edu/~bernster
