Bone marrow cells were stained with an antibody specific for the pan-B cell marker B220 and analyzed for GFP expression.
Immunoglobulin genes are assembled from their component gene segments by a series of site-specific DNA recombination reactions known as V(D)J rearrangement. The recombination reaction is targeted to individual gene segments by a cis-acting DNA sequence known as an RSS (recombination signal sequence) and by poorly understood aspects of chromatin structure which correlate with transcriptional activity of these gene-segments. In order to study the role of transcription in targeting the V(D)J recombinase, we used gene-targeting techniques to introduce a GFP cDNA into the J1 gene segment in the mouse genome. Thus, when this region undergoes transcription, the cells expressing this targeted gene turn green and can be detected and quantified by flow cytometry. To our surprise, we found that the activation of J transcription was rare and mono-allelic in developing bone marrow B cells, and that these rare, transcribed alleles were the preferred targets of the recombinase. <p>Data from Liang et al. (2004). CELL 118, 18-29.