OBTAINING STRAIN-PRODUCER OF RECOMBINANT HEXON OF BOVINE ADENOVIRUS TYPE 3
Main Article Content
Authors
K.N. Mukantayev
National center for biotechnology, Korgalzhyn road, 13/5, Astana, 010000, Kazakhstan
K.A. Tursunov
National center for biotechnology, Korgalzhyn road, 13/5, Astana, 010000, Kazakhstan
D.B. Kanayev
National center for biotechnology, Korgalzhyn road, 13/5, Astana, 010000, Kazakhstan
L. Tokhtarova
National center for biotechnology, Korgalzhyn road, 13/5, Astana, 010000, Kazakhstan
Ye. M. Ramankulov
National center for biotechnology, Korgalzhyn road, 13/5, Astana, 010000, Kazakhstan
K. K. Mukanov
National center for biotechnology, Korgalzhyn road, 13/5, Astana, 010000, Kazakhstan
Abstract
The diagnosis of bovine adenovirus is difficult due to a large number and identity of clinical symptoms with various diseases. For the diagnosis of adenoviral infection using various methods such as the isolation of the virus, in situ hybridization, enzyme-linked immunosorbent assay (ELISA), immunofluorescence, restriction analysis and polymerase chain reaction (PCR). The methods used are time consuming and require expensive equipment. In this regard, obtaining recombinant virus antigens of diagnostic importance is very important. One of the important diagnostic antigens is hexon, a group-specific protein that induces the formation of group-specific antibodies.
Under de novo conditions, the gene of the N-terminal fragment of hexon of BAV-3 with a length 503 base pairs was synthesized. A strain of E. coli BL21/pET28/rhAdv3 producing recombinant hexon of BAV-3 (rhAdv3) was obtained. The protein produced by the strain BL21/pET28/rhAdv3 contains a hexa-histidine tag and has a molecular weight of 25 kDa. An analysis of the MS/MS data using the Mascot program and the SwissProt database showed that the highest rate (Score 2033) corresponded to a fragment of hexon of BAV-3.
The gene of the fragment of the N-terminal part of the hexon of BAV-3 was synthesized. The genetic construct pET28/rhAdv3 and the microorganism strain BL21/pET28/rhAdv3 producing rhAdv3 protein were obtained. rhAdv3 has the potential to develop test systems designed to diagnose adenoviral infection of farm animals.
Keywords
gene, cloning, microorganism strain, adenovirus, recombinant protein
Article Details
References
Belak S. Ovine adenoviruses. In: Dinter Z. and Morein B. Virus infections of ruminants. Amsterdam: Elsevier Science Publishers, 1990, pp. 171-185.
Giusti A., Luini M., Benkö M., Scanziani E. Pathological and situ hybridization findings in calves experimentally infected with bovine adenovirus type 4. Dtsch Tierarztl Wochenschr, 1998, vol. 105, pp. 142-144.
Belak S. and Thoren P. Molecular diagnosis of animals’ disease: some experiences over the past decade. Expert Review of Molecular Diagnostics, 2001, Vol. 1, pp. 434-443.
Hoffmanna B., Beera M., Reid S.M., Mertens P., Oura C.A.L., van Rijnc P.A., Slomka M.J., Banks J., Brownd I.H., Alexander D.J., King D.P. A review of RT-PCR technologies used in veterinary virology and disease control: Sensitive and specific diagnosis of five livestock diseases notifiable to the World Organization for Animal Health. Veterinary Microbiology, 2009, vol. 139, pp. 1-23.
Rusvai M. and Belak S. Detection of bovine adenovirus nucleic acid sequences in nasal specimens by biotinylated DNA probes. Journal of Veterinary Medicine Series B, 1992, vol. 39, pp. 311-316.
Kiss I., Matiz K., Allard A., Wadell G., Benkö M. Detection of homologous DNA sequences in animal adenoviruses by polymerase chain reaction. Acta Veterinaria Hungarica, 1996, vol. 44, pp. 243-251.
Toogood C.I.A., Crompton J., Hay R.T. Antipeptide antisera define neutralizing epitopes on the adenovirus hexon. Journal General Virology, 1992, vol. 73, pp. 1429-1435.
Pettersson U., Wadell G. Antigenic structure of the adenoviruses. In: van Regenmortel M.H.V., Neurath. A.R. Immunochemistry of viruses. The basis for serodiagnosis and vaccines. Elsevier Science, 1985, pp. 295-323.
Sheppard M., McCoy R.J., Werner W. Genomic mapping and sequence analysis of the fowl adenovirus serotype 10 hexon gene. Journal General Virology, 1995, vol. 76, pp. 2595-2600.
Roberts M.M., White J.L., Grütter M.G., Burnett R.M. Three-dimensional structure of the adenovirus major coat protein hexon. Science, 1986, vol. 232, pp.1148-1151.
Bradford M. A rapid and sensitive method for the quantitation of microgram quantitaties of protein utilizing the principle of protein - due binding. Analytical Biochemistry, 1976, vol. 72, pp. 248-254.
Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature, 1970, VOL. 227, pp. 680-685.
Towbin P.K., Strahelin T., Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets. Proceeding National Academic Science, 1979, vol. 76, pp. 4350-4354.
Thompson, K.G., Thomson G.W., and Henry J.N. Alimentary tract manifestations of bovine adenovirus infections. Can. Vet. J, 1981, vol. 22, pp. 68-71.
Dan A., Ruzsics Z., Russell W.C., Benko M., Harrach B. Analysis of the hexon gene sequence of bovine adenovirus type 4 provides further support for a new adenovirus genus (Atadenovirus). Journal of General Virology, 1998, vol. 79, pp. 1453-1460.
Itkonen J.M., Urtti A., Bird L.E., Sarkhel S. Codon optimization and factorial screening for enhanced soluble expression of human ciliary neurotrophic factor in Escherichia coli. BMC Biotechnology, 2014, vol. 14, pp. 92-101.
Dolgova A.S., Stukolova O. A. High-fidelity PCR enzyme with DNA-binding domain facilitates de novo gene synthesis. 3 Biotech, 2017, Vol. 7, pp. 128-134.
Kalaiselvi G., Parthiban M., Narayanan M.S., Kumar S.S., Kumanan K. Rapid latex agglutination test for serodiagnosis of fowl adenovirus serotype 4 using recombinant antigen. Veterinarski arhiv, 2010, vol. 80, P.743-752.