Simultaneous isolation of pure and intact chloroplasts and mitochondria from moss as the basis for sub-cellular proteomics.
The moss Physcomitrella patens is increasingly being used as a model for plant systems biology studies. While genomic and transcriptomic resources are in place, tools and experimental conditions for proteomic studies need to be developed. In the present study we describe a rapid and efficient protocol for the simultaneous isolation of chloroplasts and mitochondria from moss protonema. Routinely, 60-100 μg mitochondrial and 3-5 mg chloroplast proteins, respectively, were obtained from 20 g fresh weight of green moss tissue. Using 14 plant compartment marker antibodies derived from seed plant and algal protein sequences, respectively, the evolutionary conservation of the compartment marker proteins in the moss was demonstrated and purity and intactness of the extracted organelles confirmed. This isolation protocol and these validated compartment markers may serve as basis for sub-cellular proteomics in P. patens and other mosses.
Lang
Erika G E
EG
Plant Biotechnology, Faculty of Biology, University of Freiburg, Schaenzlestr 1, 79104 Freiburg, Germany.
Mueller
Stefanie J
SJ
Hoernstein
Sebastian N W
SN
Porankiewicz-Asplund
Joanna
J
Vervliet-Scheebaum
Marco
M
Reski
Ralf
R
D000917:Antibody Formation; D015415:Biomarkers; D015153:Blotting, Western; D019068:Bryopsida / Q000235:genetics / Q000378:metabolism*; D002458:Cell Fractionation / Q000379:methods; D002736:Chloroplasts / Q000235:genetics / Q000378:metabolism*; D008928:Mitochondria / Q000235:genetics / Q000378:metabolism*; D010940:Plant Proteins / Q000378:metabolism*; D040901:Proteomics / Q000379:methods
D015415:Biomarkers; D010940:Plant Proteins