FIELD: biotechnology.
SUBSTANCE: said DNA aptamer binds to Shiga-toxin of type 2 and represents a single-stranded DNA AptStx2B17 with 81 nucleotides in length and molecular weight of 27 kDa. The DNA aptamer binds specifically the B-subunit of Shiga-like toxin 2 with the binding constant of 1.7·109 M-1 and primary sequence CATACGTTCGACTGCTACCTTAGCCTGGTCTTATGATGCCTGTTTGATCCTGATGGCTAATATGTGAGATGTACAGACTAG. The sequence of the DNA aptamer is selected as a result of positive selection rounds against the recombinant protein of B subunit of Shiga-toxin of type 2 based on the system of magnetic separation with separation of the target protein affinely connected with specific aptamers from the composition of the chimeric polypeptide by cleavage of the chimeric construct on site comprising protease substrate of lethal factor. The DNA aptamer is able to detect specifically the B-subunit of Shiga-like toxin 2 by PCR method in real time at a concentration of 10-11 g and to neutralise its activity towards the continuous line Vero cells at a concentration of ID50=3 mcM.
EFFECT: invention enables to detect with high efficiency the B-subunit of Shiga toxin.
4 dwg, 7 ex
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