Format

Send to

Choose Destination
Proteomics. 2015 Dec;15(23-24):4159-74. doi: 10.1002/pmic.201500246.

Purification of intact chloroplasts from marine plant Posidonia oceanica suitable for organelle proteomics.

Author information

1
Laboratory of Plant Biology and Plant Proteomics (Lab.Bio.Pro.Ve), Department of Chemistry and Chemical Technologies, Università della Calabria, Rende, Italy.
2
A.S. Diving Center Paolano, Lungomare Nord, Paola, Italy.
3
Italian National Agency for New Technologies, Energy and Sustainable Economic Development (ENEA), TRISAIA Research Center, Rotondella (Matera), Italy.
4
ALGAE - Marine Plant Ecology, Center of Marine Sciences (CCMAR), University of Algarve, Faro, Portugal.

Abstract

Posidonia oceanica is a marine angiosperm, or seagrass, adapted to grow to the underwater life from shallow waters to 50 m depth. This raises questions of how their photosynthesis adapted to the attenuation of light through the water column and leads to the assumption that biochemistry and metabolism of the chloroplast are the basis of adaptive capacity. In the present study, we described a protocol that was adapted from those optimized for terrestrial plants, to extract chloroplasts from as minimal tissue as possible. We obtained the best balance between tissue amount/intact chloroplasts yield using one leaf from one plant. After isopynic separations, the chloroplasts purity and integrity were evaluated by biochemical assay and using a proteomic approach. Chloroplast proteins were extracted from highly purified organelles and resolved by 1DE SDS-PAGE. Proteins were sequenced by nLC-ESI-IT-MS/MS of 1DE gel bands and identified against NCBInr green plant databases, Dr. Zompo database for seagrasses in a local customized dataset. The curated localization of proteins in sub-plastidial compartments (i.e. envelope, stroma and thylakoids) was retrieved in the AT_CHLORO database. This purification protocol and the validation of compartment markers may serve as basis for sub-cellular proteomics in P. oceanica and other seagrasses.

KEYWORDS:

Chloroplast purification; Marine proteomics; Plant proteomics; Seagrasses

PMID:
26444578
DOI:
10.1002/pmic.201500246
[Indexed for MEDLINE]

Supplemental Content

Full text links

Icon for Wiley
Loading ...
Support Center