Fig. 5

NAT as a selectable marker that is compatible with paromomycin and/or hygromycin B resistant genes. a Differential interference contrast (DIC) images showing wild type cells, ift54 mutant and rescued cells. Please note that ift54 did not have flagella and flagellar formation was rescued in IFT54-HA expressing cells. Bar, 10 μm. b A diagram showing construct that harbors expression cassettes of IFT54-HA and NAT. c Expression of IFT54-HA in strains harboring paromomycin resistant gene by using NAT as a selectable marker. The construct listed above was transformed into ift54, which harbors paromomycin resistant gene. The transformants were selected on agar plates supplemented with 10 µg/ml NTC. Cell lysates from randomly picked transformants were subjected to immunoblotting using the indicated antibodies. CDPK3 was used as a loading control. *Denotes non-specific bands. Wild type (WT) and ift54 cells were used as control. d Expression of IFT54-HA in strains harboring paromomycin and hygromycin B resistant genes by using NAT as a selectable marker. wdr92::WDR92-YFP strains with both paromomycin and hygromycin B resistant genes were transformed. The expression of IFT54-HA from cells grown on selection plate was examined by immunoblotting. WT and wdr92::WDR92-YFP (paro/hygro) cells were used as control