GltBD | NADPH-dependent glutamate synthase
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54.2 | 54 kDa (small subunit)
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Soluble fraction of Synechocystis PCC6803 total cell extract freshly extracted with 2x SDS-sample buffer (+ 2ME) for SDS-PAGE and denatured with 4X SDS buffer at 95°C for 5 min. Samples were separated on 10% SDS-PAGE and blotted 1h to PVDF membrane. Blot was blocked with 3 % skim milk/TBS-T, 1h/RT with agitation. Blot was incubated in the primary antibody at a dilution of 1: 2000 in TBS-T for 1h/RT. The antibody solution was decanted and the blot was washed 4 times for 10 min in TBS-T at RT with agitation. Blot was incubated in matching secondary antibody (anti-rabbit IgG horse radish peroxidase conjugated) diluted to 1:10 000 in for 1h/RT with agitation. The blot was washed as above and developed with a chemiluminescent detection reagent, following manufacture's recommendation.
Detected bands are 168 kDa (GltB, large subunit) and 54 kDa (GltD, small subunit) of NADPH-dependent glutamate synthase
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