Fluorometric analysis of the green fluorescent protein
The green fluorescent protein (eGFP) concentration in culture medium and
in TSP extracts of B. rapa rapa hairy roots was determined by
fluorometry. Undiluted spent medium samples coming from different
treatments were pipetted in pre-cooled black microtiter plates
(Microfluor 2; Thermo Fisher Scientific, Waltham, MA) as triplicates.
The fluorescence of the samples was determined at 485/527 nm using a
VICTOR2 plate reader (Perkin Elmer, Waltham, MA) at 12 nm bandwidth and
100 ms measurement time. Sample readings were compared to a standard
curve constructed with a purified GFP standard with known
concentrations. For analyzing remaining intracellular eGFP after
induction, TSP was extracted and analyzed as with the abovementioned
fluorometric analysis.