Extended Data Fig. 6: GhDIR5 and GhDIR6 expression and activity assays.
From: Dirigent gene editing of gossypol enantiomers for toxicity-depleted cotton seeds
a, Western blot of purified GhDIR5 and GhDIR6 proteins extracted from agroinfiltrated leaves of N. benthamiana and affinity purified with Ni sepharose column. Mouse anti-His antibody (TA-02, ORIGENE) was used at a dilution of 1:5000 using goat-anti-mouse IgG horseradish peroxidase conjugate (1:10000, ZB2305, ORIGENE) as secondary antibody. Molecular weight of selected bands was indicated in kDa. The experiments were repeated independently three times with similar results. b, Enantiomeric ratio of (±)-2 resulted from oxidative coupling of 1, the reaction was incubated with commercialized RvLac and GhDIR5 or GhDIR6, as indicated. Reaction without DIR was used as control. (Data are means ± s. d., n = 3 independent experiments). c, Production of total 2 increased with the amounts of GhDIR5 (0.5–5 μg in 100 μL), RvLac was used as the oxidative enzyme. 2 formed in the absence of GhDIR5 was set to 1 (means ± s. d., n = 2 or 3 independent experiments).
