Anti gfp 7 1 and 13

Two days after agroinfiltration, tobacco leaf tissue was frozen in liquid nitrogen and ground in 2× Laemmli buffer with 0.2 M dithiothreitol. The protein samples were boiled for 5 min, centrifuged to pellet leaf debris, then separated by sodium dodecylsulphate–polyacrylamide gel electrophoresis and transferred by electroblotting to a nitrocellulose membrane. The membrane was blocked in 5% skim milk then probed with anti-GFP (7.1 and 13.1; Roche, Penzberg, Germany), followed by sheep anti-mouse IgG antibody conjugated to horseradish peroxidase. Labelling was detected using the Amersham ECL Western Blotting Detection Reagent and the ImageQuant LAS 4000 CCD camera system (GE Healthcare Life Sciences, Little Chalfont, UK).