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Open Access Methodology

Isolation of dimorphic chloroplasts from the single-cell C4 species Bienertia sinuspersici

Shiu-Cheung Lung, Makoto Yanagisawa and Simon DX Chuong*

Author Affiliations

Department of Biology, University of Waterloo, 200 University Avenue West, Waterloo, Ontario N2L 3G1, Canada

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Plant Methods 2012, 8:8  doi:10.1186/1746-4811-8-8

Published: 6 March 2012

Abstract

Three terrestrial plants are known to perform C4 photosynthesis without the dual-cell system by partitioning two distinct types of chloroplasts in separate cytoplasmic compartments. We report herein a protocol for isolating the dimorphic chloroplasts from Bienertia sinuspersici. Hypo-osmotically lysed protoplasts under our defined conditions released intact compartments containing the central chloroplasts and intact vacuoles with adhering peripheral chloroplasts. Following Percoll step gradient purification both chloroplast preparations demonstrated high homogeneities as evaluated from the relative abundance of respective protein markers. This protocol will open novel research directions toward understanding the mechanism of single-cell C4 photosynthesis.

Keywords:
Bienertia sinuspersici; Chloroplast isolation; Dimorphic chloroplasts; Osmotic swelling; Photosynthesis; protoplast; Single-cell C4; Vacuole isolation