Fig. 6.
Southern blot analysis of I-SceI-induced BsdR+ colonies. (A–D) Southern blotting was performed on U2OS reporter Clones #18 (A, C) and #24 (B, D) using PstI (A, B) or HindIII (C, D) digested genomic DNA. The probe was the 700 bp GFP cDNA. See restriction maps in Fig. 5A. P: parental Clone #18 or #24 indicates the unrearranged reporter; 1–7: representative rearrangements of the reporter resulting from sister chromatid recombination events. Lanes 1–3 for Clone #18 and lanes 1–2 for Clone #24 show the most frequent LTGC products (“GFP triplication”). Lanes 4–6 for Clone #18 and lanes 3–7 for Clone #24 show aberrant patterns of expansion (see text). Note the large intensely hybridizing PstI fragment in Panel A (Clone #18), lane 4, and the correspondingly increased intensity of the 3.2 kb band in Panel C (Clone #18), lane 4. (E) Schematic of the gene amplification event in Clone #18, lane 4 (Panels A and C)—not to scale. Stars indicate positions of BsdR cassettes.