METHOD FOR DETECTING RNA OF AN ARTERITIS VIRUS AGENT IN HORSES

FIELD: biotechnology.SUBSTANCE: invention relates to the field of biotechnology. Invention is a method for detecting RNA of an agent of arteritis virus in horses, involving RNA extraction from biological material of infected individuals by a sorption method, synthesis of cDNA on an RNA matrix by single-step staging with addition of internal and positive controls of samples of a multiplex reverse transcription reaction and polymerase chain reaction with 45 cycles of amplification with real-time detection using oligonucleotide primer-specific oligonucleotide primers, a fluorescent-labeled probe and control samples, measuring accumulation of a fluorescent signal through channels of corresponding fluorescent dyes and interpreting results based on presence or absence of intersection of the fluorescence curve with a threshold line - Threshold, according to the invention, the biological material of animals is used for RNA extraction by choice: in the form of discharge from nose, eyes, faeces, blood, sperm and fragments of internal organs and tissues taken from horses infected with the arteritis virus (Equine arteritis virus) agent, wherein for internal control sample suspension of bacteriophage MS2 with concentration of 5×10copies of nucleotide sequences per 1 mcl, and for a positive control sample, a mixture of recombinant plasmid DNA containing a fragment of the genome of the horse arteritis virus (Equine arteritis virus) and a fragment of the bacteriophage MS2 genome, taken in ratio 1:1, FAM/Green for a specific signal; Cy5/Red for internal control signal, if the accumulation curves of the fluorescent signal reach 35th cycle, the result of the reaction is considered positive, and if the curves do not cross the threshold line or cross it after 35 cycles, the reaction result is negative.EFFECT: invention allows to expand functional capabilities and to conduct reliable diagnostics of the disease.1 cl, 3 dwg, 4 tbl Изобретение относится к области биотехнологии. Изобретение представляет собой способ выявления РНК возбудителя вируса артериита у лошадей, включающий выделение РНК из биологического материала инфицированных особей сорбционным методом, синтез кДНК на матрице РНК путем постановки одноэтапной с добавлением внутреннего и положительного контролей образцов мультиплексной реакции обратной транскрипции и полимеразной цепной реакции с проведением 45 циклов амплификации с детекцией в реальном времени с использованием специфичных для участка генома вирусного возбу