METHOD FOR BARCODING AMPLICONS IN THE PREPARATION OF TARGETED DNA LIBRARIES FOR MASS PARALLEL SEQUENCING Russian patent published in 2021 - IPC C12N15/00 C12Q1/68 

FIELD: molecular biology, genetic engineering.

SUBSTANCE: invention relates to the field of molecular biology and genetic engineering. A method for attaching oligonucleotide barcodes to the products of targeted DNA amplification by forming “sticky” ends of amplicons using Pfu DNA polymerase is proposed. At the first stage, the primers for targeted amplification are designed so that the 5'-end of each primer contains a given sequence of five nucleotides: 5'-TTCAG, then, after targeted amplification, the resulting amplicons are treated with Pfu DNA polymerase in the presence of the corresponding deoxyribonucleoside triphosphates (dNTP), then the barcode and one of the adapters are ligated along the formed “sticky” ends of the amplicon, while the ligation of the second adapter to the barcode occurs in the same reaction mixture using a protruding end, having a different nucleotide sequence, with the help of primers complementary to adapters, then during PCR, the reaction mixture is enriched with target sequences.

EFFECT: expanding the functionality of the method, increasing the efficiency of introducing barcodes and reducing the probability of formation of by-products when preparing targeted DNA libraries for mass parallel sequencing.

1 cl, 2 dwg, 1 tbl, 3 ex