TEST SYSTEM FOR DETECTING DNA OF LUMPY SKIN DISEASE VIRUS (LSDV) IN BIOLOGICAL MATERIAL OF ANIMALS USING A POLYMERASE CHAIN REACTION IN REAL TIME

摘要

FIELD: biotechnology.;SUBSTANCE: invention represents a test system for detecting DNA of the lumpy skin disease virus (LSDV) in animal biological material using a real-time polymerase chain reaction comprising a buffer for polymerase chain reaction, a mixture thereof, consisting of deoxynucleoside triphosphates, oligonucleotide primers and fluorescent probes specific for the genomic region of the agent of the infectious animal disease and for the internal control sample; mixture of enzymes from DNA polymerase with antibodies inhibiting enzyme activity, TAQ POLYMERASE, internal control sample in the form of suspension of bacteriophage T4 with concentration of 5×10³ phage particles per 1 mcl, negative control sample and a positive control sample, which is a mixture of recombinant plasmid DNA containing a fragment of the genome of the causative agent of the infectious animal disease and a fragment of the bacteriophage T4 genome with the nucleotide sequence: T4F: 5'-TACATATAAATCACGCAAAGC-3' – forward primer; T4R: 5'-TAGTATGGCTAATCTTATTGG-3' – reverse primer; T4P: FAM-5'-ACATTGGCACTGACCGAGTTC-3'-BHQ1 – probe, taken in volumetric ratio of 1:1, according to the invention for the internal control sample phagolysate of bacteriophage T4 is used, and for the positive control sample – fragments of genomes of native bacteriophage T4 and fragment of genome of lumpy skin disease virus (LSDV) with the following nucleotide sequence: LSDV-F 5-TTTAGGAGATAAGATTGTCAC-3' forward primer; LSDV-R 5'-GGAGATTTTATTATGAGTGGC-3' reverse primer; LSDV-P ROX-5'-GGTTAATTTTTTACCACAAATAGG-3'-BHQ2 probe.;EFFECT: invention widens functional capabilities, increases sensitivity and simplifies the process of preparing an internal control sample.;1 cl, 4 tbl