Fig. 2.

Expression Analysis of tagged MatK (A) Immunological detection of HA-tagged MatK protein. Chloroplasts isolated from transplastomic or WT lines were lysed and fractionated into membranes and soluble stroma. Fractions were separated by SDS-PAGE on a 12% gel, blotted and probed using an antiserum against the HA-epitope (Sigma). Arrow heads point to where an irrelevant marker lane has been removed from the images. RbcL = large subunit of RuBisCo detected by Ponceau staining. (B) Tagged MatK accumulation during tobacco development. Total protein was extracted from tobacco seeds or plants at different ages (41). Protein concentrations were determined and equal amounts of protein were subject to Western analysis. From top to bottom: a. Phenotypes of plants at different ages. b. Example Ponceau stain after transfer of proteins to a nylon membrane showing RbcL in green tissue, while this protein is absent in seeds and germinating seedlings. c. Exemplary chemolimunogram after detection of tagged MatK with an anti-HA antiserum in the eight tissues analyzed. d. Chemoluminescence signals from three independent Western analyses were quantified using a Lumi-Imager (Roche), analyzed with Quantity One software (Biorad) and plotted relative to the maximum signal obtained for 7d old plants (set to 1). Standard deviations are given as vertical bars.