Figure 4

Excision and re-integration of SB transposons in the progeny of double-transgenic hopper frogs. (a) GFP expression in sibling tadpoles derived from the outcross of an 8F hopper frog. Tadpoles 1 and 2 are significantly brighter than their GFP-positive siblings (tadpoles 3, 4 and 5). Tadpole 6 is a GFP-negative tadpole. Dorsal view, with anterior facing towards the right. (b) Representative data for the outcross population of 8F hopper frogs. Table includes data from breeding four F2 (8F♂54, 8F♂55, 8F♂56 and 8F♀61) and seven F3 (8F35♂A, B, C etc.) double-transgenic hoppers with wild-type frogs. The outcross progeny were scored for GFP expression and the GFP-bright progeny were either harvested for integration site analysis or raised to adulthood and outcrossed. A range of apparent remobilization activity from 0% to 0.7% was observed in individual 8F hopper frogs, with an average rate of two remobilization events per thousand GFP-positive progeny (0.2%). (c) Southern blot analysis of genomic DNA harvested from the progeny of double transgenic 8F hopper frogs. Genomic DNA was digested with Bgl II and the blot was probed with a radiolabelled GFP cDNA probe. DNA harvested from tadpoles in lanes 3, 4 and 5 have the same banding pattern as the parental pT2 βGFP 8F founder line. Lanes 1 and 2 show example of remobilization of an SB transposon. The dashed arrow indicates the change in the mobility of the transposon-harboring Bgl II fragment. Lane 6 contains DNA from GFP-negative siblings. GFP: green fluorescent protein; SB: Sleeping Beauty.