Internal IgH class switch region deletions are position-independent and enhanced by AID expression -- Data Supplement - Supporting Information -- Proceedings of the National Academy of Sciences

Supporting information for Dudley et al. (2002) Proc. Natl. Acad. Sci. USA, 10.1073/pnas.152333499

Fig. 5.
Targeting strategy for disrupting the AID gene and generation of AID^–/–/Rag-2^–/– chimeras. (A) Strategy for replacing a 3-kb region of the AID locus including exon 2 and a portion of exon 3 with the neomycin resistance gene. The WT AID locus is shown along with the AID targeting vector and the mutant allele after homologous recombination. The 3' AID probe used for detecting the targeted allele is represented by the hatched box. H, HindIII; P, Pst1; PV, Pvu1; S, Sma1; X, Xba1; TK, thymidine kinase. (B) Southern blot of DNA from WT, AID^+/–, and AID^–/– TC-1 embryonic stem cells. The DNA was digested with HindIII and probed with the 3' AID probe shown in A. (C) Flow cytometry was performed on spleen cells isolated from WT and AID^–/– Rag-1^–/– chimeras showing that AID^–/–B cells do not undergo CSR. The two FACS plots (Upper) are from day 0 spleen cells stained with B220 and IgM-specific antibodies. (Lower) The plots are from spleen cells stimulated for 4 days with anti-CD40 + IL-4 and stained with B220 and IgG1-specific antibodies.