Methods for samples preparation in proteomic research

Bodzoń‐Kułakowska, Anna; Bierczyńska-Krzysik, Anna; Dyląg, Tomasz; Drabik, Anna; Suder, Piotr; Noga, Marek; Jarzebinska, Justyna; Silberring, Jerzy

doi:10.1016/j.jchromb.2006.10.040

Cited by 214 publications

(209 citation statements)

References 405 publications

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“…When an microorganism is exposed to outer stress in forms of different bioactive compounds this could cause effects on the genomes which in turn regulates the proteins [42] (Bodson-Kulakowska et al 2007). SDS-PAGE analysis was performed to verify whether there were any differences in protein profiles between treated and untreated FOV exposed to the EON concentration.…”

Section: Discussionmentioning

confidence: 99%

Eugenol oil nanoemulsion: antifungal activity against Fusarium oxysporum f. sp. vasinfectum and phytotoxicity on cottonseeds

2015

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The current research deals with the formulation and characterization of bio-based oil-in-water nanoemulsion. The formulated eugenol oil nanoemulsion was characterized by dynamic light scattering, stability test, transmission electron microscopy and thin layer chromatography. The nanoemulsion droplets were found to have a Z-average diameter of 80 nm and TEM study reveals the spherical shape of eugenol oil nanoemulsion (EON). The size of the nanoemulsion was found to be physically stable up to more than 1-month when it was kept at room temperature (25°C). The TEM micrograph showed that the EON was spherical in shape and moderately mono or di-dispersed and was in the range of 50-110 nm. Three concentrations of the nanoformulation were used to evalute the anti-fusarium activity both in vitro and in vivo experiments. SDS-PAGE results of total protein from the Fusarium oxysporum f. sp. vasinfectum (FOV) isolate before and after treatment with eugenol oil nanoemulsion indicate that the content of extra cellular soluble small molecular proteins decreased significantly in EON-treated fungus. Light micrographs of mycelia and spores treated with EON showed the disruption of the fungal structures. The analysis of variance (ANOVA) for Fusarium wilt incidence indicated highly significant (p = 0.000) effects of concentration, genotype, and their interaction. The difference in wilt incidence between concentrations and control was not the same for each genotype, that is, the genotypes responded differently to concentrations. Effects of three EON concentration on germination percentage, and radicle length, were determined in the laboratory. One very interesting finding in the current study is that cotton genotypes was the most important factors in determining wilt incidence as it accounted for 93.18 % of the explained (model) variation. In vitro experiments were conducted to evaluate the potential phytotoxic effect of three EON concentrations. Concentration, genotype and concentration x genotype interaction were all highly significant sources of variation in seed germination; however, interaction was the first in importance as a source of variation followed by the concentration, while genotype was the least important source of variation. These results suggest the potential use of eugenol oil nanoemulsion for protecting seedcotton from Fusarium wilt infection.

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Section: Discussionmentioning

confidence: 99%

Eugenol oil nanoemulsion: antifungal activity against Fusarium oxysporum f. sp. vasinfectum and phytotoxicity on cottonseeds

2015

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“…Most frequently ion exchange, size exclusion, hydrophobic interaction, 65 partition, affinity chromatography, dye-ligand chromatography and electrophoretic methods are 66 applied [10,11]. These techniques are efficient, and often separate not only the proteins, but also 67 various protein variants: sequence variants (e.g.…”

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confidence: 99%

HPLC enrichment/isolation of proteins for post-translational modification studies from complex mixtures

Tóth

Ozohanics

Bobály

et al. 2014

Journal of Pharmaceutical and Biomedical Analysis

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10The paper describes a macroporous RP-HPLC method for separation and isolation/enrichment of 11 proteins from complex mixtures. The method is robust and efficient; using 2.1 or 4.6 mm 12 diameter columns provides sufficient material for subsequent proteomic analysis. The main 13 advantage of the method is that most protein variants are isolated in the same fraction, as 14 separation is not based on differences in isoelectric point. This is highly advantageous for 15 studying complex mixtures and post-translational modifications. Examples related to 16 glycosylation analysis are discussed in detail.

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“…Due to the consistent protein degradation observed in our results, we explored protease inhibitors as a remedial tool. Extensive usage of protease inhibitor cocktail has been demonstrated to combat protein degradation [21] which upon incorporation in our study, prevented the degradation from broad range of proteases. However, presence of 35 kDa band reflected the need to identify the candidate responsible for degradation of our protein.…”

Section: Discussionmentioning

confidence: 99%

Factors Affecting Inducible Expression of Outer Membrane Protein A (OmpA) of Shigella dysenteriae Type-1 in Lactococcus lactis Using Nisin Inducible Controlled Expression (NICE)

et al. 2015

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Potential use of Lactococcus lactis (L. lactis) as a heterologous protein expression host as well as for delivery of multiple therapeutic proteins has been investigated extensively using Nisin Inducible Controlled Expression (NICE) system. Optimum inducible expression of heterologous protein by NICE system in L. lactis depends on multiple factors. To study the unexplored role of factors affecting heterologous protein expression in L. lactis using NICE, the present study outlines the optimization of various key parameters such as inducer concentration, host's proteases and precipitating agent using Outer membrane protein A (OmpA). For efficient expression and secretion of OmpA, pSEC:OmpA vector was successfully constructed. To circumvent the troubles encountered during detection of expressed OmpA, the precipitating agent was switched from TCA to methanol.Nevertheless, detection was achieved accompanied by degraded protein products. Speculating the accountability of observed degradation at higher inducer concentration, different nisin concentrations were evaluated. Lower nisin concentrations were found desirable for optimum expression of OmpA. Consistently observed degradation was eliminated by incorporation of protease inhibitor cocktail which inhibits intracellular proteases and expression in VEL1153 (NZ9000 DhtrA) strain which inhibits extracellular protease leading to optimum expression of OmpA. Versatility and complexity of NICE system in L. lactis requires fine-tuning of target protein specific parameters for optimum expression.

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Methods for samples preparation in proteomic research

Cited by 214 publications

References 405 publications

Eugenol oil nanoemulsion: antifungal activity against Fusarium oxysporum f. sp. vasinfectum and phytotoxicity on cottonseeds

Eugenol oil nanoemulsion: antifungal activity against Fusarium oxysporum f. sp. vasinfectum and phytotoxicity on cottonseeds

HPLC enrichment/isolation of proteins for post-translational modification studies from complex mixtures

Factors Affecting Inducible Expression of Outer Membrane Protein A (OmpA) of Shigella dysenteriae Type-1 in Lactococcus lactis Using Nisin Inducible Controlled Expression (NICE)

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