Table 1.

FGD1 causes tumorigenic transformation of NIH 3T3 cells

PlasmidFocus-forming activity (%)aSoft agar growth (%)bTumorigenicity (freq)c
Vector0.10.00/6
FGD16.80.25/6
  • a NIH 3T3 cells were transfected with 3 μg of pRK5-myc plasmid or pRK5-myc-FGD1 and selected for 14 days with G418 (200 μg/ml). Protein expression was determined by Western blotting using an anti-myc antibody (9E10; Santa Cruz). Stably selected cells (103) were mixed with 106untransfected cells and then plated. Foci were scored at 7 days. Data are presented as the percentage of transfected cells that gave rise to foci. Data shown are representative of three independent assays done with duplicate plates.

  • b To assess colony formation in soft agar, each transfected cell line was seeded at 5 × 104 cells per 60-mm-diameter dish in growth medium containing 0.3% agar over a base layer of 0.6%. Colonies were counted at 21 days and expressed as a percentage of plated cells. Data shown are representative of three independent assays done in duplicate.

  • c Tumorigenic potential of the transfected cells was determined by subcutaneous inoculation into athymic nude mice (106 cells per site). For each cell line, three mice (two sites per mouse) were inoculated. Data are presented as total number of tumors per total number of sites. Mice that were negative for tumor growth were monitored for 3 months before being sacrificed. freq, frequency.