Abstract
We have produced antibodies against various membrane proteins in tobacco BY-2 cells to recognize uncharacterized organelle in the secretory system in plant cells. Possible membrane proteins were chosen using the EST information of tobacco and homologies of the EST encoding proteins with yeast and mammalian's protein markers for secretory organelles. Using recombinant proteins made from the EST sequences or synthetic peptides based on the sequence of proteins encoded by the ESTs, we made rabbit polyclonal antibodies purified by affinity chromatography. Using these antibodies, we analyzed localizations of these possible markers using density-gradient separated membrane fractions from BY-2 cells followed by immunoblotting. We also employed immunocytochemical methods at LM and EM level to localize the antigen in situ. Based on these analyses, we found that some proteins will be localized in organelles that had not characterized at the morphological level.
