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Home » Archive » 2009

TDK conference 2009

Expression of porcine circovirus capsid gene with different bacterial vectors
Kovács Eszter - year 5
SzIE, Faculty of Veterinary Science, Department of Microbiology and Infectious Diseases
Supervisors: Tamás Tuboly DVM, Attila Cságola DVM

Abstract:

Porcine circovirus type 2 (PCV2) infections are present worldwide in the swine populations and are responsible for major economical losses. PCV2 can induce a variety of clinical conditions that are usually referred to as PCV2 associated diseases (PCVADs). The first such disease described, the postweaning multisystemic wasting syndrome (PMWS), was initially observed in Canada, in 1991. By today PCV2 has already become enzootic in different geographic regions such as North America, the eastern part of Asia and Western Europe, while the infection is still spreading towards the east from the western part of Europe and towards the west from Asia. The virion has a non-enveloped icosahedral structure, with a single stranded DNA genome. Besides the pathogenic PCV type 2 viruses a non-pathogenic type (PCV1, a usual contaminant of cell cultures) is also known. The organization of the PCV1 and PCV2 genomes is identical; both contain two open reading frames (ORF). ORF1 (rep gene) is located on the positive strand and encodes the polymerase enzyme, responsible for virus replication. On the transcribed, complementary strand there is ORF2, encoding the capsid protein. Comparing PCV1 and PCV2 sequences, there is a similarity of 85 % in the ORF1 region, but only 66% identity at the ORF2 sequences. Based on this information, it is assumed that the pathogenicity of PCV2 at least partly is related to the nucleocapsid protein. At the same time the nucleocapsid, as the single structural protein of the virion, is the main immunogenic component carrying type-specific epitopes of the virus.

Several different types of vaccines and diagnostic materials are currently available on the market, but the efficiency and sensitivity of these products is sometimes questionable. There is no live virus containing vaccine available, inactivated virus vaccines and recombinant subunit vaccines are used for vaccination of piglets and sows alike. With the aid of vaccinations the clinical signs and the rate of infection can be reduced.

The aim of our research was to produce the capsid protein of PCV2, which could be used as a diagnostic material and vaccine after further purification steps. Recombinant plasmids (expression vectors) containing the ORF2 sequences were produced, and used for the transformation of competent E. coli cells. After selecting the proper clones gene expression was induced. The presence of the synthesized capsid protein was confirmed by SDS-polyacril-amide gel electrophoresis and Western-blot.



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