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Fig. 3 | Plant Methods

Fig. 3

From: Methodology: an optimized, high-yield tomato leaf chloroplast isolation and stroma extraction protocol for proteomics analyses and identification of chloroplast co-localizing proteins

Fig. 3

Silver-stained SDS–polyacrylamide gels and immunoblots with protein fractions from the chloroplast stroma isolation protocol. Total leaf proteins (homogenate), intact chloroplasts (from the 40–80% Percoll interface), and non-soluble membranes and stromal proteins released after osmotic lysis of chloroplasts are shown. a Equal amounts of protein (1 µg) from each fraction were loaded onto 12% SDS–polyacrylamide gels and silver stained. Masses of molecular weight markers are shown in kDa. b Protein blots were incubated with antisera to proteins known to reside in different chloroplast subcellular compartments and the cytosol. Due the differences in abundance of each protein in the different protein fraction and an antisera’s ability to detection tomato proteins, different amounts of protein were loaded per lane: stromal heat-shock protein 70 (HSP70; 12.5 µg), lumenal oxygen-evolving complex (OEC23; 1 µg), thylakoid membrane protein light-harvesting complex (LHCP; 1 µg), and cytosolic ribosomal protein S6 (RPS6; 50 µg). The RPS6 antisera cross-reacts with several tomato proteins. The 30-kDa RPS6 protein is solely found the total leaf homogenate; several of its cross-reacting proteins are enriched during the steps used for chloroplast stromal protein isolation. The mass (kDa) of each protein is shown

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