Identification of breast tumor mutations in BRCA1 that abolish its function in homologous DNA recombination.

ABSTRACT: Effects of breast cancer-associated gene 1 (BRCA1) missense mutations on the function of BRCA1 protein in DNA recombination have been little studied. In this report, we adapted a homology-directed recombination (HDR) assay to analyze the effects of BRCA1 mutations on this function. Using a HeLa-derived cell line with a genomically integrated recombination substrate, we expressed an endonuclease creating a double-stranded break in the substrate that the HDR assay scores by generation of green fluorescent protein-positive cells. By combining RNA interference (RNAi) that targets cellular BRCA1 mRNA with expression of RNAi-resistant BRCA1 mutants, we could effectively substitute selected point mutants to test these in the cellular recombination assay. We found that approximately 300 residues a

SUBMITTER: Ransburgh DJ 

PROVIDER: S-EPMC2943742 | biostudies-literature | 2010 Feb

REPOSITORIES: biostudies-literature