Figure 1
Reporter system to quantify CUG ambiguity in Candida albicans. (a) A recombinant gene, constructed by modifying the CaPGK gene, was used to monitor CUG ambiguity in vivo in C. albicans CAI-4 Cells. Thrombin and enterokinase sites, flanking a CUG reporter cassette, were introduced in the CaPGK in conjunction with a flag-tag epitope and a poly(his)6-tag. (b) The recombinant protein was expressed and purified to near homogeneity by nickel-agarose affinity chromatography. For high-pressure liquid chromatography-mass spectroscopy analysis, this protein was in-gel digested for 36 hours in presence of 3.0 × 10-4 U/μl of enterokinase and 3.0 × 10-5 U/μl of thrombin (Novagen).