scholarly journals Generation and characterization of the VP1 recombinant protein of the chicken anemia virus

2019 ◽  
Vol 2019 (2) ◽  
pp. 12-20
Author(s):  
Михаил Грудинин ◽  
Mihail Grudinin ◽  
Андрей Комиссаров ◽  
Andrey Komissarov ◽  
Алина Гусейнова ◽  
...  
Keyword(s):  
Recombinant Protein ◽  
Recombinant Proteins ◽  
Polyclonal Antibodies ◽  
Chicken Anemia Virus ◽  
Glutathione S Transferase ◽  
Adenoviral Infection ◽  
Metal Affinity ◽  
Infectious Laryngotracheitis ◽  
Potential Component ◽  
Anemia Virus

The aim of this study is to obtain a VP1 recombinant protein of the chicken anemia virus, capable of specifically detecting antibodies in the blood sera of sick chickens. Materials and methods. Cloning of a fragment of the VP1 gene of an infectious anemia virus of chickens was performed in the expression plasmids pET15b and pGEX-3T in the context of reading the polyhistidine sequence and glutathione-S-transferase, respectively. The recombinant proteins 6HIS-ΔVP1 and GST-ΔVP1 expressed in E. coli Rosetta (DE3) strains were purified by metal affinity chromatography. Amino acid sequence of recombinant proteins was confirmed by mass spectrometry. The specificity of the interaction of recombinant proteins with polyclonal antibodies was determined by ELISA. The ability of the recombinant 6HIS-ΔVP1 protein to detect antibodies in field and blood sera of SPF chickens was evaluated by indirect ELISA. To control the specificity of the antigen, the immune sera of birds for viruses of infectious bronchitis, Newcastle disease, infectious laryngotracheitis, adenoviral infection, Mycoplasma gallisepticum, and negative serum of chickens were used. Results. The recombinant VP1 protein of chicken anemia virus containing a polyhistidine tag (6HIS-∆VP1) was obtained. It was shown that this recombinant protein is able to specifically detect antibodies in the blood sera of sick chickens. Conclusion. The obtained recombinant protein 6HIS-ΔVP1 can be used to detect antibodies to the chicken anemia virus in the serum of sick chickens, and can also be considered as a potential component of vaccines against this virus.

scholarly journals Multiple Microfermentor Battery: a Versatile Tool for Use with Automated Parallel Cultures of Microorganisms Producing Recombinant Proteins and for Optimization of Cultivation Protocols

2006 ◽  
Vol 72 (8) ◽  
pp. 5225-5231 ◽  
Author(s):  
Emmanuel Frachon ◽  
Vincent Bondet ◽  
Hélène Munier-Lehmann ◽  
Jacques Bellalou
Keyword(s):  
Recombinant Protein ◽  
Recombinant Proteins ◽  
Protein Production ◽  
Batch Cultures ◽  
E Coli ◽  
Working Volume ◽  
Versatile Tool ◽  
Labview Program ◽  
Medium Formulation ◽  
Conventional Medium

ABSTRACT A multiple microfermentor battery was designed for high-throughput recombinant protein production in Escherichia coli. This novel system comprises eight aerated glass reactors with a working volume of 80 ml and a moving external optical sensor for measuring optical densities at 600 nm (OD600) ranging from 0.05 to 100 online. Each reactor can be fitted with miniature probes to monitor temperature, dissolved oxygen (DO), and pH. Independent temperature regulation for each vessel is obtained with heating/cooling Peltier devices. Data from pH, DO, and turbidity sensors are collected on a FieldPoint (National Instruments) I/O interface and are processed and recorded by a LabVIEW program on a personal computer, which enables feedback control of the culture parameters. A high-density medium formulation was designed, which enabled us to grow E. coli to OD600 up to 100 in batch cultures with oxygen-enriched aeration. Accordingly, the biomass and the amount of recombinant protein produced in a 70-ml culture were at least equivalent to the biomass and the amount of recombinant protein obtained in a Fernbach flask with 1 liter of conventional medium. Thus, the microfermentor battery appears to be well suited for automated parallel cultures and process optimization, such as that needed for structural genomics projects.

scholarly journals Inhibition of Fungal Appressorium Formation by Pepper (Capsicum annuum) Esterase

2001 ◽  
Vol 14 (1) ◽  
pp. 80-85 ◽  
Author(s):  
Young Soon Kim ◽  
Hyun Hwa Lee ◽  
Moon Kyung Ko ◽  
Chae Eun Song ◽  
Cheol-Yong Bae ◽  
...  
Keyword(s):  
Recombinant Protein ◽  
Capsicum Annuum ◽  
Magnaporthe Grisea ◽  
Dependent Manner ◽  
Appressorium Formation ◽  
Glutathione S Transferase ◽  
Porcine Liver ◽  
Esterase Gene ◽  
Dose Dependent ◽  
Dependent Signaling Pathway

A pepper esterase gene (PepEST) that is highly expressed during an incompatible interaction between pepper (Capsicum annuum) and the anthracnose fungus Colletotrichum gloeosporioides has been previously cloned. Glutathione-S-transferase-tagged recombinant PepEST protein expressed in Escherichia coli showed substrate specificity for p-nitrophenyl esters. Inoculation of compatible unripe pepper fruits with C. gloeosporioides spores amended with the recombinant protein did not cause anthracnose symptoms on the fruit. The recombinant protein has no fungicidal activity, but it significantly inhibits appressorium formation of the anthracnose fungus in a dose-dependent manner. An esterase from porcine liver also inhibited appressorium formation, and the recombinant protein inhibited appressorium formation in the rice blast fungus, Magnaporthe grisea. Inhibition of appressorium formation in M. grisea by the recombinant protein was reversible by treatment with cyclic AMP (cAMP) or 1,16-hexadecanediol. The results suggest that the recombinant protein regulates appressorium formation by modulating the cAMP-dependent signaling pathway in this fungus. Taken together, the PepEST esterase activity can inhibit appressorium formation of C. gloeosporioides, which may result in protection of the unripe fruit against the fungus.

Production of cocktail of polyclonal antibodies using bacterial expressed recombinant protein for multiple virus detection

2014 ◽  
Vol 196 ◽  
pp. 7-14 ◽  
Author(s):  
Reetika Kapoor ◽  
Bikash Mandal ◽  
Prabir Kumar Paul ◽  
Phaneendra Chigurupati ◽  
Rakesh Kumar Jain
Keyword(s):  
Recombinant Protein ◽  
Polyclonal Antibodies ◽  
Virus Detection ◽  
Multiple Virus

scholarly journals Performance comparison between broilers positive and negative for antibodies against the chicken anemia virus

2003 ◽  
Vol 34 ◽  
pp. 88-89
Author(s):  
Lauricio Librelotto Rubin ◽  
Luiz Antônio Faccenda de Ávila ◽  
Andréa Machado Leal Ribeiro ◽  
Vera Wald ◽  
Cláudio Wageck Canal
Keyword(s):  
Performance Comparison ◽  
Chicken Anemia Virus ◽  
Anemia Virus

Immobilized metal-ion affinity systems for recovery and structure–function studies of proteins at molecular, supramolecular, and cellular levels

2010 ◽  
Vol 82 (1) ◽  
pp. 39-55 ◽  
Author(s):  
Rajasekar R. Prasanna ◽  
Mookambeswaran A. Vijayalakshmi
Keyword(s):  
Capillary Electrophoresis ◽  
Recombinant Proteins ◽  
Gel Electrophoresis ◽  
Metal Ion ◽  
Immobilized Metal Affinity Chromatography ◽  
Metal Affinity ◽  
Purification Technique ◽  
Affinity Gel Electrophoresis ◽  
Insight Into ◽  
Histidine Tag

Immobilized metal-ion affinity (IMA) adsorption is a collective term that is used to include all kinds of adsorptions where the metal ion serves as the characteristic and most essential part of adsorption center. Of all the IMA techniques, immobilized metal-affinity chromatography (IMAC) has been gaining popularity as the choice of purification technique for proteins. IMAC represents a separation technique that is primarily useful for proteins with natural surface exposed-histidine residues and for recombinant proteins with engineered histidine tag. This review also gives insight into other nonchromatographic applications of IMA adsorption such as immobilized metal-ion affinity gel electrophoresis (IMAGE), immobilized metal-ion affinity capillary electrophoresis (IMACE), and immobilized metal-ion affinity partitioning (IMAP).

scholarly journals Positive and Negative Regulation of Chicken Anemia Virus Transcription

2005 ◽  
Vol 79 (5) ◽  
pp. 2859-2868 ◽  
Author(s):  
Myrna M. Miller ◽  
Keith W. Jarosinski ◽  
Karel A. Schat
Keyword(s):  
Estrogen Receptor ◽  
Nuclear Receptor ◽  
Protein Translation ◽  
Chicken Anemia Virus ◽  
Egfp Expression ◽  
Start Site ◽  
Response Element ◽  
Nuclear Receptor Superfamily ◽  
Promoter Construct ◽  
Anemia Virus

ABSTRACT Chicken anemia virus (CAV) is a small circular single-stranded DNA virus with a single promoter-enhancer region containing four consensus cyclic AMP response element sequences (AGCTCA), which are similar to the estrogen response element (ERE) consensus half-sites (A)GGTCA. These sequences are arranged as direct repeats, an arrangement that can be recognized by members of the nuclear receptor superfamily. Transient-transfection assays which use a short CAV promoter construct that ended at the transcription start site and drive expression of enhanced green fluorescent protein (EGFP) showed high basal activity in DF-1, LMH, LMH/2A, and primary theca and granulosa cells. The estrogen receptor-enhanced cell line, LMH/2A, had significantly greater expression than LMH cells, and this expression was significantly increased with estrogen treatment. A long promoter construct which included GGTCA-like sequences downstream of the first CAV protein translation start site was found to have significantly less EGFP expression in DF-1 cells than the short promoter, which was largely due to decreased RNA transcription. DNA-protein binding assays indicated that proteins recognizing a consensus ERE palindrome also bind GGTCA-like sequences in the CAV promoter. Estrogen receptor and other members of the nuclear receptor superfamily may provide a mechanism to regulate CAV activity in situations of low virus copy number.

Genome sequencing analysis of Brazilian chicken anemia virus isolates that lack MSB-1 cell culture tropism

2007 ◽  
Vol 30 (2) ◽  
pp. 81-96 ◽  
Author(s):  
Eliana Ottati Nogueira ◽  
Antonio J Piantino Ferreira ◽  
Rodrigo Martins Soares ◽  
Edison Luiz Durigon ◽  
Simaia Lazzarin ◽  
...  
Keyword(s):  
Cell Culture ◽  
Genome Sequencing ◽  
Chicken Anemia Virus ◽  
Sequencing Analysis ◽  
Anemia Virus ◽  
Virus Isolates
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