METHOD OF IDENTIFYING DNA TISSUE OF JAPANESE MACKEREL (SCOMBER JAPONICUS) IN FISH PRODUCTS, MEAT AND BONE FISH MEAL AND FEED USING REAL-TIME POLYMERASE CHAIN REACTION Russian patent published in 2024 - IPC C12Q1/68 

FIELD: veterinary microbiology.

SUBSTANCE: method of identifying DNA tissue of Japanese mackerel (Scomber japonicus) in fish products and in meat and bone fish meal contained in feed is described. DNA is isolated from animal tissue using the sorption method. A polymerase chain reaction with fluorescent detection is carried out with 45 cycles of amplification in real time using oligonucleotide primers, probes, fluorescent dyes specific to the animal DNA genome region: for a specific signal for the animal — JOE/Yellow and Cy5/Red — for an internal control sample in the form of a suspension of bacteriophage T4 with a concentration of 5×10³ phage particles per 1 µl, positive control sample in the form of a mixture containing fragments of animal genomes and bacteriophage T4 with the following nucleotide sequence: T4F TACATATAAATCACGCAAAGC is direct primer, T4R TAGTATGGCTAATCTTATTGG is reverse primer, T4P CY5 ACATTGGCACTGACCGAGTTC is probe taken in a 1:1 ratio. The accumulation of fluorescent signals through the channels of the corresponding fluorescent dyes is measured. The results are interpreted based on the presence or absence of intersection of the fluorescence curve with the threshold line. Thus, if the accumulation curves of the fluorescent signal appear before the 35th cycle, the reaction result is considered positive, and if the curves do not cross the threshold line or cross it after the 35th cycle, the reaction result is negative. According to the invention, DNA is isolated from the tissue of Japanese mackerel (Scomber japonicus) and a fragment of the genome of tissue of Japanese mackerel (Scomber japonicus) with the following nucleotide sequence used for a positive control sample: Sj-F ATAGCCCTCGCCCTAACTCT primer, Sj-R AGGGCTCATTTGTGTCCAGG primer, Sj-Z R6G ATCAGCGGAGGGTAGGTGTA BHQ1 probe.

EFFECT: invention can be used to expand the functionality and increase the accuracy of identifying the species of fish tissue.

1 cl, 5 tbl, 1 ex