Figure 4.

V-DJ rearrangements on the WT Igh^b allele of V_HΔ^a/WT^b mice. (A) Igh locus. Probe C detects a sequence deleted upon V-DJ joining. Probe D detects a sequence retained on both alleles in all cells. (B) Assay for V-DJ rearrangement on the Igh^a and Igh^b alleles in WT^a/WT^b and V_HΔ^a/WT^b mice. Genomic DNA extracted from liver and from sorted IgM^a−b+ spleen cells (WT^a/WT^b; left) and IgM^a+b− spleen cells (V_HΔ^a/WT^b; right). DNAs were digested with BamHI and hybridized with probes C (top) and D (bottom). (C) Quantitative analysis (ImageQuant) of blot shown in B. A repeat experiment was done using liver and spleen cells from V_HΔ^a/WT^b animals. Band intensities in liver were set to 100%. Normalization was to probe D for WT^a/WT^b mice; normalization was to the Igh^a fragment in V_HΔ^a/WT^b mice. Error bars show SD. (D) Igh locus maps indicating PCR primers and probes for detecting V-DJ rearrangements on Igh^b allele. 5′ primers specific for individual V_H gene families (V_H family primer) were used in combination with a 3′ primer (J_H4 primer), which was present on the Igh^b allele but missing on both the V_HEμ^a and V_HΔ^a alleles. The representative blot demonstrates allele-specific amplification. H₂O, no DNA template control.