Evaluation of protein extraction methods for enhanced proteomic analysis of tomato leaves and roots

Vilhena, Milca Bartz; Franco, Mônica Regina; Schmidt, Daiana; Carvalho, Gabriela Ferreira; Azevedo, Ricardo Antunes

doi:10.1590/0001-3765201520150116

Cited by 14 publications

(9 citation statements)

References 32 publications

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“…1) is in accordance with the results reported by Gomez-Martınez (2012). Future studies should perhaps employ more detailed analysis of Pearl Millet storage proteins by using 2D-PAGE, as has been done for a wide range of crops species (Arruda et al 2013, Vilhena et al 2015, Schmidt et al 2016.…”

Section: Discussionmentioning

confidence: 99%

Growth, Yield and Grain Nutritional Quality in Three Brazilian Pearl Millets (Pennisetum americanum L.) with African or Indian origins

Médici

Gonçalves

Fonseca

et al. 2018

An. Acad. Bras. Ciênc.

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In this study, we are presenting recommendations to the best agricultural use as well as for plant breeding of three millet cultivars namely ENA1 and ENA2, which have African origin, and BRS1501 originally from India. These cultivars were evaluated for growth, yield and grain quality traits. The morphological traits evaluated in this study indicated that the African genotypes ENA1 and ENA2 are better than the Indian genotype BRS1501 for no-till farming or to produce forage with 15% of crude protein at flowering and at harvest to produce stover (around 7% of crude protein content) for livestock feeding. The BRS1501 cultivar exhibited the highest values for total crude protein, albumins and prolamins, phytate and mineral contents in grains. ENA1 and ENA2 exhibited the highest values of globulin and glutelin contents. The electrophoretic patterns for storage proteins were similar across the three millets cultivars, except for a higher intensity of two glutelin bands with 21 and 24 kDa in BRS1501. Together, the results allow us to recommend BRS1501 for grain production and ENA1 and ENA2 for biomass production.

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Section: Discussionmentioning

confidence: 99%

Growth, Yield and Grain Nutritional Quality in Three Brazilian Pearl Millets (Pennisetum americanum L.) with African or Indian origins

Médici

Gonçalves

Fonseca

et al. 2018

An. Acad. Bras. Ciênc.

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“…However, it does not necessarily mean that TCA cannot extract proteins with these functions because these functional proteins may be present in the proteins co-extracted by the two or three methods. At present, many protein extraction method studies are only focus on one proteomic technique, the 2-DE or high-throughput proteomic techniques [31–33]. The use of proteins extracted by the same method for both 2-DE and LC-MS/MS studies can avoid false differential abundant proteins associated with different extraction methods, allowing identification of a higher number of proteins with various isoforms [10, 34].…”

Section: Discussionmentioning

confidence: 99%

An improved protein extraction method applied to cotton leaves is compatible with 2-DE and LC-MS

et al. 2019

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Background Two-dimensional electrophoresis (2-DE) and liquid chromatography-tandem mass spectrometry (LC-MS/MS) are widely used in plant proteomics research. However, these two techniques cannot be simultaneously satisfied by traditional protein extraction methods when investigate cotton leaf proteome. Results Here, we evaluated the efficiency of three different protein extraction methods for 2-DE and LC-MS/MS analyses of total proteins obtained from cotton leaves. The protein yield of the borax/PVPP/phenol (BPP) method (0.14%) was significantly lower than the yields of the trichloroacetic acid/acetone (TCA) precipitation method (1.42%) and optimized TCA combined with BPP (TCA-B) method (0.47%). The BPP method was failed to get a clear 2-DE electrophoretogram. Fifty pairs of protein spots were randomly selected from the 2-DE gels of TCA- and TCA-B-extracted proteins for identification by MALDI TOF/TOF, and the results of 42 pairs were consistent. High-throughput proteomic analysis showed that 6339, 9282 and 9697 unique proteins were identified from the total cotton leaf proteins extracted by the TCA, BPP and TCA-B methods, respectively. Gene Ontology (GO) analysis revealed that the proteins specifically identified by TCA method were primarily distributed in the plasma membrane, while BPP and TCA-B methods specific proteins distributed in the cytosol, indicating the sub-cellular preference of different protein extraction methods. Further, ATP-dependent zinc metalloprotease FTSH 8 could be observed in the 2-DE gels of TCA and TCA-B methods, and could only be detected in the LC-MS/MS results of the BPP and TCA-B methods, showing that TCA-B method might be the optimized choice for both 2-DE and LC-MS/MS. Conclusion Our data provided an improved TCA-B method for protein extraction that is compatible with 2-DE and LC-MS/MS for cotton leaves and similar plant tissues which is rich in polysaccharides and polyphenols. Electronic supplementary material The online version of this article (10.1186/s12864-019-5658-5) contains supplementary material, which is available to authorized users.

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“…The pellet fraction at the bottom with enriched cysts was collected for protein extraction. A trichloroacetic acid/acetone precipitation with phenol extraction method was used to isolate total proteins from enriched cysts, as previously described [37]. The concentrations of extracted proteins were measured by the Bradford protein assay before use.…”

Section: Methodsmentioning

confidence: 99%

Activation of chronic toxoplasmosis by transportation stress in a mouse model

et al. 2016

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Toxoplasma gondiiis an obligate intracellular parasite infecting 25% of the world population and enormous number of animals. It can exist in two forms in intermediate hosts: the fast replicating tachyzoites responsible for acute infection and the slowly replicating bradyzoites responsible for life-long chronic infection. The interconversion between tachyzoites and bradyzoites plays critical roles in the transmission and pathogenesis of T. gondii. However, the molecular mechanisms that govern the interconversion are largely unknown. In this study, we established a chronic infection model in mice and examined the impact of transportation stress on the status of chronic infection. Our results demonstrated that, treating chronically infected mice with conditions mimicking transportation stress reduced the levels of several key cytokines that restrict the infection at chronic stage. Increased expression of the tachyzoite specific gene SAG1 (surface antigen 1) was detected in brain cysts of stress treated mice, indicating activation and conversion of bradyzoites to tachyzoites. Using this model, we identified fifteen toxoplasmic proteins that had significant abundance changes during stress induced cysts reactivation. These proteins serve as a basis for further investigation of the mechanisms governing bradyzoite conversion.

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Evaluation of protein extraction methods for enhanced proteomic analysis of tomato leaves and roots

Cited by 14 publications

References 32 publications

Growth, Yield and Grain Nutritional Quality in Three Brazilian Pearl Millets (Pennisetum americanum L.) with African or Indian origins

Growth, Yield and Grain Nutritional Quality in Three Brazilian Pearl Millets (Pennisetum americanum L.) with African or Indian origins

An improved protein extraction method applied to cotton leaves is compatible with 2-DE and LC-MS

Activation of chronic toxoplasmosis by transportation stress in a mouse model

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