Isolating intact chloroplasts from small Arabidopsis samples for proteomic studies.
We have established a method for the isolation of chloroplasts from Arabidopsis thaliana that allows proteomic studies in the context of biotic stress with small amounts of starting material. Employing a 50% Percoll layer to separate crude filtrates, the required leaf material was reduced to 2-3g, yielding more than 300 microg of chloroplast proteins. The quality of this fraction was confirmed by immunological, enzymatic, and gel-based assays. This protocol provides intact chloroplasts from Arabidopsis plants with a high degree of integrity and purity as well as sufficient protein recovery, thereby enabling studies of plant-herbivore or plant-pathogen interactions.
Kley
Jeannette
J
Max Planck Institute for Chemical Ecology, Jena, Germany.
Heil
Martin
M
Muck
Alexander
A
Svatos
Ales
A
Boland
Wilhelm
W
D017360:Arabidopsis / Q000737:chemistry / Q000166:cytology* / Q000201:enzymology / Q000378:metabolism; D015415:Biomarkers / Q000378:metabolism; D002736:Chloroplasts / Q000737:chemistry / Q000201:enzymology / Q000378:metabolism; D003584:Cytological Techniques / Q000379:methods*; D003600:Cytosol / Q000378:metabolism; D008928:Mitochondria / Q000378:metabolism; D018515:Plant Leaves / Q000737:chemistry; D010940:Plant Proteins / Q000032:analysis / Q000737:chemistry; D040901:Proteomics; D011786:Quality Control; D015203:Reproducibility of Results; D012995:Solubility
D015415:Biomarkers; D010940:Plant Proteins