Fig. 1. Generation of Ret-TGM mice. (A) Homologous recombination between Ret gene (top) and the targeting vector (second line) resulted in insertion of TGM and floxed-Tn5-neo (third line). Tn5-neo was successively removed in mice to generate Ret-TGM allele (bottom). Arrowheads indicate loxP sites. (B) Southern blot analysis for RetTGM mutation using genomic DNA from RW-4 ES cells (control), homologous recombinant ES clone (clone 108), and tails of wild-type (+/+), heterozygous (TGM/+) and homozygous (TGM/TGM) mice. The DNA samples were digested with NcoI and hybridized with radiolabeled probe, indicated in Fig. 1A. (C) Representative pictures of TGM expression in the nervous system. Signals obtained by RET immunohistochemistry (left) completely overlaps that of GFP fluorescent signal provided by TGM (middle) in trigeminal ganglion of newborn RetTGM/+ mice (merged image, right). (D) No RET protein expression is detected in spinal motoneurons (left) that are identified by GFP fluorescence (right) in RetTGM/TGM newborn mice. Abbreviations: K, KpnI; N, NcoI. Scale bar: 55 µm in C; 100 µm in D.
