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. 2000 Jun 27;97(14):7975–7980. doi: 10.1073/pnas.130190597

Figure 4.

Figure 4

Analysis of DJβRPF, DJβRC, and DJβRD rearrangement in T cells. (A) Schematic of DJβRPF, DJβRC, and DJβRD miniloci. In DJβRPF, the germ-line Dβ1/Jβ1.1 gene segments and intervening sequence of TCRβPF has been replaced by a preassembled DJβ1.1 rearrangement. In DJβRC, the position of DJβ1.1 and Jβ1.2 and their flanking RSSs have been exchanged. In DJβRD, the position of the coding region of DJβ1.1 and Jβ1.2 has been exchanged with the RSSs remaining in their native positions. The Jβ1.2 12-RSS (shaded triangle), Dβ1 12-RSS (open triangle), and Vβ14 23-RSS (dotted open triangle) are indicated. (B) BglII-digested genomic DNA isolated from kidney (K) and ES cells transfected with DJβRPF or thymocytes (T) and B cells (B) from a mouse with the DJβRPF minilocus was subjected to Southern blot analysis by using probe A (Fig. 1). Shown are the expected size bands from the nonrearranged endogenous TCRβ locus (end) and nonrearranged (GL) or VDJ/VJ-rearranged DJβRPF minilocus. The 9- and 6-kb markers are indicated. (C) The PA and PB primers were used to PCR thymocyte DNA from wild-type non-miniloci-containing mice (T) or mice that have the TCRβPF (−1), DJβRPF (−8, −19), DJβRC (−24, −35), or DJβRD (−11, −13) miniloci. Analyses shown are from mice generated from independently derived ES cells. Genomic DNA was digested with BglII before PCR; PCR products were subjected to Southern blot analysis by using probe A. Vβ rearrangement to the Jβ1.2 RSS (shaded triangle) yields a 700-bp PCR product from DJβRC and 600-bp PCR products from DJβRPF and DJβRD.