Bulletin "Veterinary biotechnology"

Veterynarna biotehnologija – Veterinary biotechnology, 2021, 39, 88-95 [in Ukrainian]. https://doi.org/10.31073/vet_biotech39-08

MUZYKINA L.M., e-mail: This email address is being protected from spambots. You need JavaScript enabled to view it., MOLOZHANOVA A.V., е-mail: This email address is being protected from spambots. You need JavaScript enabled to view it., SYDORENKO T.V., е-mail: This email address is being protected from spambots. You need JavaScript enabled to view it., TARASOV O.A., е-mail: This email address is being protected from spambots. You need JavaScript enabled to view it., MEZHENSKA N.A., e-mail: This email address is being protected from spambots. You need JavaScript enabled to view it., SYTIUK M.P., e-mail: This email address is being protected from spambots. You need JavaScript enabled to view it.

Institute of Veterinary Medicine NAAS

ASF VIRUS DNA ISOLATION USING DIFFERENT KITS FOR NUCLEIC ACID ISOLATION IN CASE OF “ASFPCR-RT” TEST KIT APPLICATION

Introduction. African swine fever (ASF) is a highly contagious viral disease of domestic and wild pigs that causes huge economic loss to Ukrainian pig production. There are no treatment and specific prevention of the disease, so the determining factor of the veterinary and sanitary measures to control ASF is rapid in time diagnostic approaches. The stage of sample preparation and DNA isolation is important in the polymerase chain reaction (PCR) test for African swine fever diagnostics.

The goal of the work. To establish the efficiency of ASF virus DNA isolation by sorption, precipitating and column isolation kits for ASF PCR- RT diagnostic test.

Materials and methods. ASF virus-containing pig spleens (5 samples) of and ASF-negative blood of pigs (5 samples) were used for test validation. For amplification “Diagnostic kit ASF PCR-RT for the diagnosis of African swine fever by real-time PCR” was used. Three nucleic acid extraction kit: “DNA-sorb-B”, “RIBO-prep”, “IndiSpin Pathogen Kit” were used to obtain nucleic acids from the samples.

Results of research and discussion. According to the results of the research, it was established that nucleic acid extraction kits “DNA-sorb-B” and “RIBO-prep” allowed to get enough DNA for 10 tested samples according to concentration specified by the test kit, but “IndiSpin Pathogen Kit” failed for one positive porcine spleen sample, which could be due to high concentration of nucleic acid in sample.

Conclusion and prospects for future research. It is necessary to apply the recommendations of the manufacturer of the nucleic acid extraction kit “IndiSpin Pathogen Kit”, which recommend to use 10fold dilution of the spleen samples for DNA isolation not more then 5-10 mg of sample.

Keywords: DNA isolation, ASF virus, pigs, spleen, whole blood, PCR.

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