FIG. 2.

Culture conditions for inducible centrin RNAi. A. Anticentrin staining of methanol-fixed nitRNAi1 cells 2 and 6 days after transfer to NH₄Cl-free medium. Arrowheads (day 2), cells with short NBBCs. Bar, 5 μm. B. The % of nitRNAi1 cells displaying wild-type centrin fibers on different days after transfer to ammonium-free medium. Inset: Cells from day 4 lacking NBBCs but retaining dCFs. C. Time course showing the % of nitRNAi2 cells displaying a wild-type phenotype after return to NH₄Cl medium. Inset: Cells examined at intermediate times (day 4) frequently displayed single NBBCs and a punctate staining on the cell nucleus but lacked dCFs. D. nitRNAi1 cells initially maintained in ammonium-containing medium with a light-dark (L/D) cycle of 14 hours/10 hours were transferred to the medium and conditions indicated. The % of cells with a wild-type phenotype was determined by anticentrin indirect immunofluorescence at different time points. a, KNO₃ as a nitrogen source and constant light at 25°C (▪) or a light-dark cycle of 14 h/10 h at 22°C (○). b, KNO₃ as a nitrogen source with a light-dark cycle of 14 h/10 h in medium supplemented with 1 μm Na₂MoO₄ (○) or Na₂WO₄ (•). c, NH₄Cl as a nitrogen source and constant light (▪) or a light-dark cycle of 14 h/10 h (○).