METHOD FOR DETECTING TOXIN GENES BY MULTIPLEX PCR WITH SUBSEQUENT SEQUENCING Russian patent published in 2021 - IPC G01N33/53 C12N15/00 

FIELD: medicine; microbiology; molecular genetics.

SUBSTANCE: invention relates to medicine, namely to the field of microbiology, molecular genetics, it allows detecting toxin genes of bacterial origin in environmental samples, food, clinical samples. A multiplex panel of oligonucleotides is created for amplification of toxin genes. To do this, oligonucleotides are selected for targeted amplification based on the following criteria: the length of oligonucleotides should be at least 17 nucleotides; the size of the amplified fragments is from 200 to 600 nucleotides; primers should not form dimers and secondary structures; primers for different targets should not form dimers and secondary structures and should be assembled into a multiplex panel. Then, based on the above criteria, a multiplex panel of oligonucleotides is developed for amplification of a fragment of the diphtheria toxin gene, the streptococcal toxin gene of group A, the cholera toxin gene of subunit A, the cholera toxin gene of subunit B, the botulinum toxin gene of type A, the botulinum toxin gene of type B, the botulinum toxin gene of type C1, the botulinum toxin gene of type D, the botulinum toxin gene of type E, the botulinum toxin gene of type F, the botulinum toxin gene of type G, the tetanus toxin gene. Their primers are obtained. Then, using the selected oligonucleotides, the corresponding sections of the genome of Corynebacterium diphtheriae, Streptococcus pyogenes, Vibrio cholerae, Clostridium botulinum and Clostridium tetani microorganisms are amplified.

EFFECT: provided is possibility of detecting toxin genes.

2 cl, 1 dwg, 2 tbl