scholarly journals Biomanufacturing of Tomato-Derived Nanovesicles

Foods ◽  
2020 ◽  
Vol 9 (12) ◽  
pp. 1852
Author(s):  
Ramesh Bokka ◽  
Anna Paulina Ramos ◽  
Immacolata Fiume ◽  
Mauro Manno ◽  
Samuele Raccosta ◽  
...  
Keyword(s):  
Buoyant Density ◽  
Morphological Characteristics ◽  
Sucrose Density Gradient ◽  
Size Exclusion ◽  
High Yield ◽  
Plant Resources ◽  
Edible Plant ◽  
Solanum Lycopersicum L ◽  
In Silico Studies ◽  
Shock Proteins

Micro- and nano-sized vesicles (MVs and NVs, respectively) from edible plant resources are gaining increasing interest as green, sustainable, and biocompatible materials for the development of next-generation delivery vectors. The isolation of vesicles from complex plant matrix is a significant challenge considering the trade-off between yield and purity. Here, we used differential ultracentrifugation (dUC) for the bulk production of MVs and NVs from tomato (Solanum lycopersicum L.) fruit and analyzed their physical and morphological characteristics and biocargo profiles. The protein and phospholipid cargo shared considerable similarities between MVs and NVs. Phosphatidic acid was the most abundant phospholipid identified in NVs and MVs. The bulk vesicle isolates were further purified using sucrose density gradient ultracentrifugation (gUC) or size-exclusion chromatography (SEC). We showed that SEC using gravity column efficiently removed co-purifying matrix components including proteins and small molecular species. dUC/SEC yielded a high yield of purified vesicles in terms of number of particles (2.6 × 1015 particles) and protein quantities (6.9 ± 1.5 mg) per kilogram of tomato. dUC/gUC method separated two vesicle populations on the basis of buoyant density. Proteomics and in silico studies of the SEC-purified MVs and NVs support the presence of different intra- and extracellular vesicles with highly abundant lipoxygenase (LOX), ATPases, and heat shock proteins (HSPs), as well as a set of proteins that overlaps with that previously reported in tomato chromoplast.

scholarly journals Identification of Tomato Infecting Viruses That Co-Isolate with Nanovesicles Using a Combined Proteomics and Electron-Microscopic Approach

Nanomaterials ◽  
2021 ◽  
Vol 11 (8) ◽  
pp. 1922
Author(s):  
Ramila Mammadova ◽  
Immacolata Fiume ◽  
Ramesh Bokka ◽  
Veronika Kralj-Iglič ◽  
Darja Božič ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Mosaic Virus ◽  
Protein Identification ◽  
Plant Viruses ◽  
Size Exclusion ◽  
Tomato Mosaic Virus ◽  
High Yield ◽  
Plant Resources ◽  
Electron Microscopic ◽  
Viral Particles

Plant-derived nanovesicles (NVs) have attracted interest due to their anti-inflammatory, anticancer and antioxidative properties and their efficient uptake by human intestinal epithelial cells. Previously we showed that tomato (Solanum lycopersicum L.) fruit is one of the interesting plant resources from which NVs can be obtained at a high yield. In the course of the isolation of NVs from different batches of tomatoes, using the established differential ultracentrifugation or size-exclusion chromatography methods, we occasionally observed the co-isolation of viral particles. Density gradient ultracentrifugation (gUC), using sucrose or iodixanol gradient materials, turned out to be efficient in the separation of NVs from the viral particles. We applied cryogenic transmission electron microscopy (cryo-TEM), scanning electron microscopy (SEM) for the morphological assessment and LC–MS/MS-based proteomics for the protein identification of the gradient fractions. Cryo-TEM showed that a low-density gUC fraction was enriched in membrane-enclosed NVs, while the high-density fractions were rich in rod-shaped objects. Mass spectrometry–based proteomic analysis identified capsid proteins of tomato brown rugose fruit virus, tomato mosaic virus and tomato mottle mosaic virus. In another batch of tomatoes, we isolated tomato spotted wilt virus, potato virus Y and southern tomato virus in the vesicle sample. Our results show the frequent co-isolation of plant viruses with NVs and the utility of the combination of cryo-TEM, SEM and proteomics in the detection of possible viral contamination.

EVALUATION OF THE COMMON HOP VARIETY SAMPLES ACCORDING TO THE PHENOLOGICAL AND MORPHOLOGICAL FEATURES

2018 ◽  
pp. 40-42 ◽  
Author(s):  
А. А. Fadeev ◽  
Z. А. Nikonova
Keyword(s):  
Morphological Characteristics ◽  
Economic Importance ◽  
Morphological Features ◽  
High Yield ◽  
Early Maturity ◽  
Foreign Countries ◽  
Humulus Lupulus L ◽  
The Common ◽  
Regions Of Russia ◽  
Vegetative Season

The results of study of the 12 year cycle of studies on the only in Russia collection of hops ordinary (Humulus lupulus L.), which contains 250 samples from different regions of Russia and 17 foreign countries. The number of process varieties, composition and origin, it is unique and corresponds to world level. A collection of accessions of hops is a population of female plants with a set of phenological, morphological and economic importance of signs. In the article, the estimation of the collectible varieties of hops at different ripeness groups according to phenological and morphological characteristics according to the method of test for distinctness, uniformity and stability. As the result of the research the Common Hop (Humuluslupulus) sorts were classified in accordance with their maturity time as early maturity (less than 100 days) – 10%, middle-early (101…110 days) – 14, middle duration (111…120 days) – 40, middle-late (121…130 days) – 10% and slow-maturing (more than 130 days) – 26%. Each group has a phenotypic and morphologies features. The early maturity, middle-early and middle duration varieties with vegetative season approximately 120 days are more adaptive to the conditions of the Chuvashia and central part of the Russia and provide obtaining high yield of the hop cones.

Evaluation of the Efficiency of Protein A Affinity Chromatography to Purify a Monoclonal Antibody for Cancer Treatment and its Purity Analysis

2020 ◽  
Vol 7 (2) ◽  
pp. 121-133
Author(s):  
Ayesha Akhtar ◽  
Shivakumar Arumugam ◽  
Shoaib Alam
Keyword(s):  
Monoclonal Antibody ◽  
Monoclonal Antibodies ◽  
Affinity Chromatography ◽  
Protein A ◽  
Exchange Chromatography ◽  
Size Exclusion ◽  
High Yield ◽  
Staphylococcal Protein A ◽  
Purification Step ◽  
Purity Analysis

Background:: Protein A affinity chromatography is often employed as the most crucial purification step for monoclonal antibodies to achieve high yield with purity and throughput requirements. Introduction:: Protein A, also known as Staphylococcal protein A (SPA) is found in the cell wall of the bacteria staphylococcus aureus. It is one of the first discovered immunoglobulin binding molecules and has been extensively studied since the past few decades. The efficiency of Protein A affinity chromatography to purify a recombinant monoclonal antibody in a cell culture sample has been evaluated, which removes 99.0% of feed stream impurities. Materials and Method:: We have systematically evaluated the purification performance by using a battery of analytical methods SDS-PAGE (non-reduced and reduced sample), Cation Exchange Chromatography (CEX), Size-exclusion chromatography (SEC), and Reversed phased-Reduced Chromatography for a CHO-derived monoclonal antibody. Results and Discussion:: The analytical test was conducted to determine the impurity parameter, Host Cell Contaminating Proteins (HCP). It was evaluated to be 0.015ng/ml after the purification step; while initially, it was found to be 24.431ng/ml. Conclusion:: The tests showed a distinct decrease in the level of different impurities after the chromatography step. It can be concluded that Protein A chromatography is an efficient step in the purification of monoclonal antibodies.

scholarly journals Isolation of HDL by sequential flotation ultracentrifugation followed by size exclusion chromatography reveals size-based enrichment of HDL-associated proteins

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Jack Jingyuan Zheng ◽  
Joanne K. Agus ◽  
Brian V. Hong ◽  
Xinyu Tang ◽  
Christopher H. Rhodes ◽  
...  
Keyword(s):  
Structural Integrity ◽  
Polyacrylamide Gel Electrophoresis ◽  
Cholesterol Acyltransferase ◽  
Density Lipoprotein ◽  
Size Exclusion ◽  
High Yield ◽  
Phospholipid Transfer ◽  
Exclusion Chromatography ◽  
Associated Proteins ◽  
Alpha 1 Antitrypsin

AbstractHigh-density lipoprotein (HDL) particles have multiple beneficial and cardioprotective roles, yet our understanding of their full structural and functional repertoire is limited due to challenges in separating HDL particles from contaminating plasma proteins and other lipid-carrying particles that overlap HDL in size and/or density. Here we describe a method for isolating HDL particles using a combination of sequential flotation density ultracentrifugation and fast protein liquid chromatography with a size exclusion column. Purity was visualized by polyacrylamide gel electrophoresis and verified by proteomics, while size and structural integrity were confirmed by transmission electron microscopy. This HDL isolation method can be used to isolate a high yield of purified HDL from a low starting plasma volume for functional analyses. This method also enables investigators to select their specific HDL fraction of interest: from the least inclusive but highest purity HDL fraction eluting in the middle of the HDL peak, to pooling all of the fractions to capture the breadth of HDL particles in the original plasma sample. We show that certain proteins such as lecithin cholesterol acyltransferase (LCAT), phospholipid transfer protein (PLTP), and clusterin (CLUS) are enriched in large HDL particles whereas proteins such as alpha-2HS-glycoprotein (A2HSG), alpha-1 antitrypsin (A1AT), and vitamin D binding protein (VDBP) are enriched or found exclusively in small HDL particles.

scholarly journals Molecular heterogeneity of mouse duodenal alkaline phosphatase. Association of lipids and peptides

1974 ◽  
Vol 141 (1) ◽  
pp. 93-101 ◽  
Author(s):  
P. R. V. Nayudu ◽  
Fraser B. Hercus
Keyword(s):  
Alkaline Phosphatase ◽  
Gradient Centrifugation ◽  
Polyacrylamide Gel Electrophoresis ◽  
Buoyant Density ◽  
Sucrose Density Gradient ◽  
Molar Ratio ◽  
Molecular Forms ◽  
Partial Specific Volume ◽  
Molecular Heterogeneity ◽  
Sucrose Density Gradient Centrifugation

Polyacrylamide-gel electrophoresis and Bio-Gel P-300 molecular-sieve chromatography of mouse duodenal alkaline phosphatase demonstrates its molecular heterogeneity, which, in a kinetic sense, is manifest also in the differential relative velocities of the heterogeneous forms of the enzyme with two substrates, phenylphosphate and β-glycerophosphate. Different treatments that eliminate most of the electrophoretic and chromatographic variability of the enzyme also decrease the velocities with both substrates so that the molar ratio of hydrolysis of one substrate relative to the other is also altered to a low but stable value. Concomitant with these changes, lipids and peptides are dissociated from the enzyme. The lipids are tentatively identified as a sterol and phospholipids. The peptides have an average composition of four to six amino acids and appear to be strongly electropositive. The conditions of dissociation suggest that their binding to the enzyme is non-covalent and predominantly based on hydrophobic and ionic bonding. The concept of lipid and peptide association would suggest prima facie differential molecular weights as a factor in the observed electrophoretic and chromatographic heterogeneity. However, the molecular forms of the enzyme with differences in elution volume equivalent to more than one-half the void volume of the Bio-Gel P-300 column, or even enzyme fractions dissociated from the lipids and peptides compared with undissociated portions, do not show any differences in sedimentation on sucrose-density-gradient centrifugation. This may be because the alterations in molecular weight owing to binding of small molecules are too small to be detected by this method. Alternatively, since lipids are involved, the binding may alter the partial specific volume in such a way that the buoyant density is not significantly altered.

scholarly journals Improved downstream process for the production of plasmid DNA for gene therapy.

2005 ◽  
Vol 52 (3) ◽  
pp. 703-711 ◽  
Author(s):  
Jochen Urthaler ◽  
Wolfgang Buchinger ◽  
Roman Necina
Keyword(s):  
Gene Therapy ◽  
Plasmid Dna ◽  
Viral Vectors ◽  
Continuous System ◽  
Size Exclusion ◽  
High Yield ◽  
Downstream Process ◽  
Naked Dna ◽  
Supercoiled Form ◽  
Exclusion Chromatography

Gene therapy and genetic vaccines promise to revolutionize the treatment of inherited and acquired diseases. Since viral vectors are generally associated with numerous disadvantages when applied to humans, the administration of naked DNA, or DNA packed into lipo- or polyplexes emerge as viable alternatives. To satisfy the increasing demand for pharmaceutical grade plasmids we developed a novel economic downstream process which overcomes the bottlenecks of common lab-scale techniques and meets all regulatory requirements. After cell lysis by an in-house developed gentle, automated continuous system the sequence of hydrophobic interaction, anion exchange and size exclusion chromatography guarantees the separation of impurities as well as undesired plasmid isoforms. After the consecutive chromatography steps, adjustment of concentration and final filtration are carried out. The final process was proven to be generally applicable and can be used from early clinical phases to market-supply. It is scaleable and free of animal-derived substances, detergents (except lysis) and organic solvents. The process delivers high-purity plasmid DNA of homogeneities up to 98% supercoiled form at a high yield in any desired final buffer.

scholarly journals Diversity of wild edible plants traditionally used by the Galo tribe of Indian Eastern Himalayan state of Arunachal Pradesh

2020 ◽  
Vol 7 (4) ◽  
Author(s):  
Padma Raj Gajurel ◽  
Tajum Doni
Keyword(s):  
Relative Frequency ◽  
Vital Role ◽  
Leafy Vegetables ◽  
Wild Edible Plants ◽  
Wild Plants ◽  
Wild Plant ◽  
Edible Plants ◽  
Arunachal Pradesh ◽  
Plant Resources ◽  
Edible Plant

Wild edible plants are found very useful in the fulfilment of food and nutritional requirements. Because of the availibity and cultural preference, the consumption of these plants among the tribes is high. To find out the diversity, utilisation pattern and sociocultural importance of the wild plants, a study was conducted in the state of Arunachal Pradesh selecting the Galo tribe, and accordingly the wild edible plants consumed are documented here. Data were collected through extensive field surveys and interviews with the community in the selected 12 villages in Upper Subansiri and West Siang districts of Arunachal Pradesh. Overall, 125 wild edible plant species under 99 genera and 54 families are reported. These species are consumed mostly as leafy vegetables, fruits, medicine, spices and condiments and as a substitute to food grains. The Urticaceae with ten species is the most utilised family followed by Asteraceae, Moraceae and Lamiaceae with at least five species in each. Herbs with 47 species were found to be the most dominant growth form followed by trees with 44 species. Based on parts used leaves with 66 species were recorded to be the most used plant parts followed by fruits. The highest edibility index of 50 % was reported in Solanum americanum. The analysis of relative frequency of citation revealed that total 78 species exhibits more than 0.50 relative frequency of citation value with highest value in Pouzolzia hirta (0.95). It has been found that the wild plant resources play a vital role in the socio-economic aspects of the Galo tribe.

scholarly journals ASSESSMENT OF THE CORRELATION OF INTERRELATIONS OF MORPHOLOGICAL CHARACTERISTICS OF THE GRAIN SORGHUM

2020 ◽  
pp. 38-42
Author(s):  
Victoria Igorevna Starchak ◽  
Valery Ivanovich Zhuzhukin ◽  
Ekaterina Aleksandrovna Zhuk ◽  
Vera Valeryevna Bychkova
Keyword(s):  
Plant Height ◽  
Flag Leaf ◽  
Morphological Characteristics ◽  
Correlation Coefficients ◽  
Coefficient Matrix ◽  
Grain Sorghum ◽  
High Yield ◽  
Total Contribution ◽  
Morphological And Physiological Traits ◽  
Research And Design

In modern conditions, the need for cultivation of grain sorghum in regions with insufficient moisture is determined by its high yield and grain quality. For practical breeding, with the objective of assessing linkages 19 morphological and physiological traits model of the population including 15 varieties and 2 promising lines of grain sorghum that is created in the Russian Research and Design Technological Institute of Sorghum and Maize. The biochemical composition of the grain of the objects of research is presented graphically. A different degree of variation in the characteristics of grain sorghum was revealed: very strong (V> 40,0%) - productive bushiness, number of grains from 1 panicle; strong (20,0% <V <40,0%) - the width of the panicle, the extension of the panicle leg, the thickness of the upper internode, the area of the flag leaf, the area of the fourth on top of the leaf, the mass of the grain from 1 panicle; weak (V <10,0%) - the height of the plants after 30 days, the height of the plants during maturation. The group with an average degree of variation (10.0% <V <20.0%) includes all other signs measured in the experiment. Factor analysis was used to optimize, interpret the calculated matrix of correlation coefficients. In the analysis of the correlation coefficient matrix, hypothetical factors with a contribution of more than 5% to the accumulated variance are calculated. The first hypothetical factor is determined by the high effects of morphophysiological features. The second factor is largely due to the contribution of plant height 30 days after germination and the extension of the panicle legs. Plant height at maturity, weight of 1000 grains make the greatest contribution to the third factor. The width of the panicle makes the greatest contribution to the accumulated dispersion of the fourth factor. The fifth factor is determined by the effects of plant height at the beginning and end of vegetation, as well as the total contribution of all studied features.

Is LM7 a new human retrovirus or a mycoplasma virion?

1995 ◽  
Vol 1 (2) ◽  
pp. 88-94 ◽  
Author(s):  
P Froussard
Keyword(s):  
Cell Culture ◽  
Reverse Transcription ◽  
Buoyant Density ◽  
Sucrose Density Gradient ◽  
Enveloped Virus ◽  
Polymerase Chain ◽  
Serological Data ◽  
Culture Supernatants ◽  
Viral Genomic Rna ◽  
Human Retrovirus

A putative retrovirus called LM7 was recently isolated from a patient with MS. This retrovirus was detected in LM7 and LM711 cultured human leptomeningeal cells. In the present work, nucleic adds from LM711 cell culture supernatants were purified and subjected to avian myeloblastosis viral (AMV) reverse transcriptase and to random polymerase chain reaction (rPCR) in order to characterize the genomic material of LM7 virions. Analysis of reverse transcription products allowed the detection of an approximately 14 kb ribonucleic acid in all LM711 cell culture supernatants. However, sequencing of rPCR-amplified molecules as well as RNA blotting data showed essentially that all tested cells producing LM7 particles were infected with mycoplasma. Moreover, purification of LM7 particles onto a linear sucrose density gradient established that the 14 kb nucleic acid was always associated with the 1.19–1.21 g ml-1 sucrose fractions, which are known to correspond to the buoyant density of mycoplasma. In addition, no viral genomic RNA was detected in the 1.17 g ml-1 sucrose fraction containing the low reverse transcription activity. These results strongly suggest that microscopic images and serological data could be related to mycoplasma and/or to a virion associated with the bacteria. The LM7 particle might be a new and additional enveloped virus able to infect Mycoplasma hyorhinis hosts. Thus, for instance, it would be presumptuous to assert, with our current understanding, that the LM7 virion is one of the causal agents of MS in humans.

scholarly journals Genetic variability and trait relationship in cherry tomato (Solanum lycopersicum L. var. cerasiforme (Dunnal) A. Gray)

2013 ◽  
Vol 41 (2) ◽  
pp. 163-167 ◽  
Author(s):  
MS Islam ◽  
HC Mohanta ◽  
MR Ismail ◽  
MY Rafii ◽  
MA Malek
Keyword(s):  
Genetic Variability ◽  
Solanum Lycopersicum ◽  
Fruit Weight ◽  
Fruit Yield ◽  
High Yield ◽  
Genetic Advance ◽  
Cherry Tomato ◽  
Solanum Lycopersicum L ◽  
Wide Range ◽  
Additive Gene

Nine traits of 11 cherry tomato (Solanum lycopersicum L.) var. cerasiforme (Dunal) A. Gray) inbred lines exhibited a wide range of genetic variability. High geno- and phenotypic coefficients of variation were obtained for individual fruit weight (68.16 and 74.23%, respectively) followed by number of fruits/plant (58.8 and 68.34%, respectively). High estimates of heritability, genetic advance and genotypic coefficient of variation for the traits of individual fruit weight, number of fruits and clusters/plant were controlled by additive gene action indicating the possibility of selection to improve these characters. Fruit yield/plant showed low heritability along with low genetic advance and did not show significant and positive correlation with the remaining characters. It indicates that improvement of high yield through selection is difficult, rather hybridization can be effective for improving the fruit yield/plant. Among the lines, CH154 produced the highest number of fruits/plant (291) and highest fruit yield (1.89 kg/plant and 63.4 t/ha) and can be selected for cultivation under Bangladesh condition. DOI: http://dx.doi.org/10.3329/bjb.v41i2.13443 Bangladesh J. Bot. 41(2): 163-167, 2012 (December)

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