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. 2008 Jun 27;36(13):4317–4326. doi: 10.1093/nar/gkn383

Figure 1.

Figure 1.

Analysis of retrotransposition in mutants of classical nuclear transport proteins. (A) A schematic of the Ty1 retrotransposition assay (38). Yeast cells are transformed with the URA3-marked pAR100 (His- cells) or pSD600 (His+ cells) test plasmid and patched onto SC ura− glu plates. Patches are then replica plated to SC ura− gal plates for 3 days at 25°C and sequentially replica plated to (1) SC ura− glu; (2) YPD; (3) 5-FOA; and (4) SC his− glu for pAR100 or YEPD + G418 for pSD600. (B) Transposition levels in nuclear protein import mutants. gsp1-1, ntf2-1, srp1-5, rsl1-L63A, and Δsxm1 mutant cells were transformed with the appropriate test plasmids and replica plated to selection medium. The final selection plates are YEPD + G418 (gsp1-1, ntf2-1, rsl1-L63A, and Δsxm1) and SC his− glu (srp1-55). As controls, each plate also contained wildtype cells either expressing (+) or lacking (−) the appropriate test plasmid. (C) Transport mutants carrying the Ty1 test plasmid grow approximately as well as wildtype cells on selective media. gsp1-1, ntf2-1, rsl1-L63A and Δsxm1 cells were transformed with the pSD600 (G418) test plasmid while srp1-55 cells were transformed with the pAR100 (His) test plasmid. Cell numbers were equalized, cultures were serially diluted 10-fold, and spotted onto either YEPD + G418 plates (top) or SC-his (bottom) glu plates. Plates were then grown at 25°C for 3 days. Wildtype cells either carrying the appropriate Ty1 test plasmid (+) or an empty vector (−) served as positive and negative controls, respectively.