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In vitro V(D)J recombination: Signal joint formation

Cortes, Patricia and Weis-Garcia, Frances and Misulovin, Ziva and Nussenzweig, Andre and Lai, Jiann-Shiun and Li, Gloria and Nussenzweig, Michel C. and Baltimore, David (1996) In vitro V(D)J recombination: Signal joint formation. Proceedings of the National Academy of Sciences of the United States of America, 93 (24). pp. 14008-14013. ISSN 0027-8424. http://resolver.caltech.edu/CaltechAUTHORS:CORpnas96

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Abstract

The first step of V(D)J recombination, specific cleavage at the recombination signal sequence (RSS), can be carried out by the recombination activating proteins RAG1 and RAG2. In vivo, the cleaved coding and signal ends must be rejoined to generate functional antigen receptors and maintain chromosomal integrity. We have investigated signal joint formation using deletion and inversion substrates in a cell free system. RAG1 and RAG2 alone or in combination were unable to generate signal joints. However, RAG1 and RAG2 complemented with nuclear extracts were able to recombine an extrachromosomal substrate and form precise signal joints. The in vitro reaction resembled authentic V(D)J recombination in being Ku-antigen-dependent.


Item Type:Article
Additional Information:© 1996 The National Academy of Sciences. Contributed by David Baltimore, August 26, 1996. We thank Dr. David Schatz for critical review of this manuscript and also for providing the DNA substrate to test the 12/23 rule, Bruce Meyer for providing the pEBG plasmid, and Juan Carcamo and Dennis Sawchuk for comments on the manuscript and very helpful discussions. P.C. was supported by a fellowship from the Irvington Institute and is now supported by the Lymphoma Research Foundation of America. J.-C.L. is supported by a fellowship from the Irvington Institute. This work was supported by grants from the National Institutes of Health to D.B. and M.C.N. M.C.N. is an associate investigator in the Howard Hughes Medical Institute and D.B. is an American Cancer Society Research Professor.
Subject Keywords:recombination activating protein RAG1, recombination activating protein RAG2, broken DNA-molecules, expression vector, mouse thymocytes, RAG-1, region, genes, rearrangement, proteins, breaks, ends
Record Number:CaltechAUTHORS:CORpnas96
Persistent URL:http://resolver.caltech.edu/CaltechAUTHORS:CORpnas96
Alternative URL:http://www.pnas.org/cgi/content/abstract/93/24/14008
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:576
Collection:CaltechAUTHORS
Deposited By: Tony Diaz
Deposited On:25 Aug 2005
Last Modified:26 Dec 2012 08:40

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