I need to extract proteins from subcellular fractions (membranes and/or vesicles, nuclei, cytoplasm, others). Does anyone know (easy) protocols for that?
Additional information: I work on filamentous fungi, plant protocols generally work well.
Hello Sabine,
I am not sure about it, but I would suggest you to look for the kit if any commercially available. I think there might be some kit available for the protein extraction.
I guess, you might have looked for it by now.
Wish you all the best.
You can separate subcellular fractions easily by centrifugation using Percoll gradients.
Good luck!
If you do not mind having all the different fractions together you first have to centrifuge at 10,000 g then precipitate the supernatant at 100,000g what is pelleted is the mix of them.
http://www.sciencedirect.com/science/article/pii/S0167779907001813
This paper contains several references that might help you
Many thanks for all suggestions. Ill try to figure out which protocol to use.
Ici il y a un protocol pour extaire les organelles, après j'ai fait l’extraction de l'ADN. La technique pour les organelles est la même, il te faudra trouver comment extraire les proteines. Infante H. Diógenes, Weissbach Arthur, (1990), Organellar DNA replication in N. tabacum cultured cells, Plant Mol. Biol. 14:891-897.
See: Analysis of Saccharomyces cerevisiae Asp3p Protein Sorting During Nitrogen Starvation In: III Bienal de Pesquisa da Fundação Oswaldo Cruz, 2002, Rio de Janeiro. Vice-Presidência de Pesquisa e Desenvolvimento Tecnológico, 2002. v.1. p.86 - 87
or Asparaginase II GFP Fusion as a Tool for Studying the Secretion of the Enzyme Under Nitrogen Starvation. Brazilian Journal of Microbiology. , v.34, p.373 - 377, 2003.
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