Extraction of plant proteins for two‐dimensional electrophoresis

Granier, Fabienne

doi:10.1002/elps.1150091106

Cited by 190 publications

(117 citation statements)

References 49 publications

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“…To examine total proteins by two-dimensional gel electrophoresis (48), we extracted proteins by using modifications of published methods (20,21). Cells harvested from a 50-ml culture were suspended in 2 ml of prechilled extraction buffer (50 mM Tris-HCl, pH 7.5, 5 mM MgCl 2 ).…”

Section: Methodsmentioning

confidence: 99%

Identification and Characterization of a Developmentally Regulated Protein, EshA, Required for Sporogenic Hyphal Branches in Streptomyces griseus

et al. 2001

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To identify sporulation-specific proteins that might serve as targets of developmental regulatory factors in Streptomyces, we examined total proteins of Streptomyces griseus by two-dimensional gel electrophoresis. Among five proteins that were present at high levels during sporulation but absent from vegetative cells, two of the proteins, P3 and P4, were absent from developmental mutants that undergo aberrant morphogenesis. The deduced amino acid sequence of the gene that encodes P3 (EshA) showed extensive similarity to proteins from mycobacteria and a cyanobacterium, Synechococcus, that are abundant during nutritional stress but whose functions are unknown. Uniquely among these proteins, EshA contains a cyclic nucleotide-binding domain, suggesting that the activity of EshA may be modulated by a cyclic nucleotide. The eshA gene was strongly expressed from a single transcription start site only during sporulation, and accumulation of the eshA transcript depended on a developmental gene, bldA. During submerged sporulation, a null mutant strain that produced no EshA could not extend sporogenic hyphae from new branch points but instead accelerated septation and spore maturation at the preexisting vegetative filaments. These results indicated that EshA is required for the growth of sporogenic hyphae and localization of septation and spore maturation but not for spore viability.Streptomyces is a gram-positive bacterium that undergoes morphological differentiation. In a nutritionally favorable condition, Streptomyces spores germinate to give rise to vegetative hyphae, which are characterized by filamentous, multigenomic cells called mycelia. Sporulation begins by the growth of aerial hyphae (or sporogenic hyphae when the sporulation process is induced in liquid culture; 34), which subsequently undergo septation to form chains of unicellular spores. We previously identified two types of sporogenic hyphae during sporulation of Streptomyces griseus (22): we suggest the term "distal" sporogenic hyphae for those that grow from the tips of preexisting vegetative filaments and the term "proximal" sporogenic hyphae for those that grow de novo within the vegetative hyphae at the onset of sporulation. Since both vegetative growth and spore formation are inherently polar processes similar to those observed in yeast and filamentous fungi (18), formation of sporogenic hyphae in Streptomyces is likely to require proteins that direct the location of sporogenic hyphae, establishment of polarity, and emergence of the sporogenic hyphae.To understand the molecular details of Streptomyces sporulation, researchers have identified developmental genes primarily by complementation of nonsporulating mutants. Most of the sporulation-specific genes that have been characterized appear to encode regulatory functions. Some, such as bldA, which encodes tRNA (38), are required for the production of aerial hyphae (35, 43) and antibiotics (3, 34, 43). Genes that specifically regulate spore formation include the whi genes, so named because mutations in ...

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Section: Methodsmentioning

confidence: 99%

Identification and Characterization of a Developmentally Regulated Protein, EshA, Required for Sporogenic Hyphal Branches in Streptomyces griseus

et al. 2001

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“…The extraction of plant tissues for the detection of LTP was performed according to Granier (1988). Polyclonal antibodies produced against purified LTP of Capsicum spp seeds were prepared as described by Diz (2007).…”

Section: Western Blottingmentioning

confidence: 99%

Peroxidase is involved in Pepper yellow mosaic virus resistance in Capsicum baccatum var. pendulum

Gonçalves

Rodrigues²,

Diz³

et al. 2013

Genet. Mol. Res.

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ABSTRACT. Pathogenesis-related proteins (PRs) are among the defense mechanisms of plants that work as an important barrier to the development of pathogens. These proteins are classified into 17 families according to their amino acid sequences, serology, and/or biological or enzyme activity. The present study aimed to identify PRs associated with the pathosystem of Capsicum baccatum var. pendulum: Pepper yellow mosaic virus (PepYMV). Forty-five-day-old plants from accession UENF 1624, previously identified as resistant to PepYMV, were inoculated with the virus. Control and infected leaves were collected for analysis after 24, 48, 72, and 96 h. The inoculated and control plants were grown in cages covered with anti-aphid screens. Proteins were extracted from leaf tissue and the presence of β-1,3-glucanase, chitinase, peroxidase, and lipid transport protein was verified.

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“…Leaf samples were taken from the pooled samples of leaves from 20 independent rice seedlings grown under PEG-created water stress at 0.5 MPa for 0, 48, and 96 h. Total proteins were extracted from 200 mg of leaf samples after being ground in liquid nitrogen and precipitated with trichloroacetic acid/ acetone according to Granier (1988) with slight modifications. The protein concentrations were determined as described by Lowry et al (1951).…”

Section: Gelc-ms/ms Shotgun Proteomicsmentioning

confidence: 99%

Physiological and comparative proteomic analyses of Thai jasmine rice and two check cultivars in response to drought stress

Maksup

Roytrakul

Supaibulwatana

2012

Journal of Plant Interactions

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To understand the molecular responses of Thai jasmine rice (Oryza sativa L. ssp. indica cv. KDML105) under drought, proteomes of KDML105 cultivars and two check cultivars, namely NSG19 (tolerant) and IR20 (sensitive) were investigated. The effect of PEG6000 and mannitol for creating osmotic stress ( 0.1 MPa to 2.1 MPa) was tested in KDML105 before applied to the three cultivars. The leaf proteomes were analyzed by GeLC-MS/MS shotgun proteomics. Among 623 proteins identified, 53 proteins showed significant difference in protein expressions, which could be hierarchically classified into three clusters of proteins up-regulated in IR20, NSG19, or in KDML105. Coronatine-insensitive 1 protein that thought to be involved in stomatal closing was prominently observed in NSG19, regarding to its physiology of rapid stomatal closing and highest stability of photosystem II. WD-40 repeat protein which involves in cell death was induced in IR20. On the other hand, H-protein promoter binding factor-2a remarkably increased in KDML105.

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Extraction of plant proteins for two‐dimensional electrophoresis

Cited by 190 publications

References 49 publications

Identification and Characterization of a Developmentally Regulated Protein, EshA, Required for Sporogenic Hyphal Branches in Streptomyces griseus

Identification and Characterization of a Developmentally Regulated Protein, EshA, Required for Sporogenic Hyphal Branches in Streptomyces griseus

Peroxidase is involved in Pepper yellow mosaic virus resistance in Capsicum baccatum var. pendulum

Physiological and comparative proteomic analyses of Thai jasmine rice and two check cultivars in response to drought stress

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