Initiation of V(D)J recombination in a cell-free system
January 1995
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Cells performing V(D)J recombination make specific cuts in DNA at recombination signal sequences. Here, we show that nuclear extracts of pre-B cell lines carry out this specific cleavage. The products of cleavage are the same as found previously in thymocytes: full-length, blunt, 5'-phosphorylated signal ends, and covalently sealed (hairpin) coding ends. A complete signal sequence is required. Recombinant RAG1 protein greatly increases activity and complements an inactive extract from a RAG1 (-/-) pre-B cell line. When the extracts are fractionated, cleavage activity correlates with the presence of RAG2 protein. These results suggest that RAG1 and RAG2 are components of the V(D)J recombinase.
- Molecular Sequence Data
- animals
- DNA/genetics
- DNA Primers/genetics
- *DNA-Binding Proteins
- cell line
- *Homeodomain Proteins
- *Recombination, Genetic
- base sequence
- Receptors, Antigen, T-Cell/*genetics
- B-Lymphocytes/immunology
- *Genes, Immunoglobulin
- cell-free system
- signal
- rag 1 protein
- protein
- cleavage
- recombination
- reaction
- rag 2 protein
- activity
- rag 1
- coding
- product
- sequence
- 12- signal
- rag 2
- coding ends
- figure
- extract
- 12-
- recombination signal sequences
- fraction
