BRI1 (Protein BRASSINOSTEROID INSENSITIVE 1) is a receptor which binds brassinolide and has a dual specificity kinase activity acting on both serine/threonine- and tyrosine-containing substrates. Involved in a signaling cascade including expression of li
Antibody was tested on bri1-1 and bri1-5 mutants. Bri1-1 is a point mutation in the kinase domain that renders the protein non-functional and plants compensate for that by over-accumulating the non-functional receptor. Bri1-5 is a mutant in the extracellular domain and the bri1-5 protein is retained in the ER. The bri1-5 plants contain less protein than the wild type and show an intermediate brassinosteroid deficient phenotype. Also BRI1-5 migrates higher than wild type BRI1 in SDS-PAGE, because it carries ER-type high mannose N-glycans. For IP: 15 µl GFP-trap beads was used for 200 mg plant material to precipitate GFP-tagged protein followed by detection with Co-IPed BRI1 on Western with 1:5000 diluted anti-BRI1 antibody.Protein extraction has to be done efficiently as this step is crucial, recommended material to buffer ratio: 15 µl/µg or less.
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