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. 2017 Jan 31;7:41720. doi: 10.1038/srep41720

Figure 6. Determination of the translation start site of the threonine substitution mutant.

Figure 6

(a) HeLa FUT2 cells were transfected with the myc-tagged versions of A transferase (A, Δ19, Δ25, etc.), A-M1,20,26,43,53,69T-myc mutant (T mutant), and the same construct with an in-frame STOP codon at 5′ of the first threonine (A-STOP-M1,20,26,43,53,69T-myc denoted as STOP-T mutant in the figure). Cell extracts were subjected to SDS-PAGE (10% polyacrylamide) and blotted onto PVDF membrane. Signals were obtained using a monoclonal anti-myc antibody followed by horseradish peroxidase-linked anti-mouse IgG antibody incubation and enhanced chemiluminescence reagent. Signals were detected on chemiluminescence sensitive film. The numbers indicate the sizes and positions of molecular weight markers in kDa. (b) The N-terminal sequence chromatograms obtained by the Procise 494 analyzer are shown for A-STOP-M1,20,26,43,53,69T-myc. In each cycle the major amino acid detected has been marked by an arrowhead and its one-letter symbol.