GENOMIC ENGINEERING Russian patent published in 2022 - IPC C12N15/09 C12N15/90 C12N5/10 A61K48/00 

FIELD: biotechnology; cellular engineering.

SUBSTANCE: present invention relates to biotechnology and cellular engineering, in particular to a method for modifying a target DNA in a stem cell expressing Cas enzyme from a CRISPR type II system, which forms a colocalization complex with a guide RNA, complementary to the target DNA, and which cleaves the target DNA in a site-specific manner, and to a stem cell for RNA-directed Cas-based genome editing, which is not a human germ line cell or a human embryonic cell. To implement the specified method, the guide RNA, complementary to the target DNA, is first introduced into the specified stem cell, wherein RNA and Cas enzyme are elements of the colocalization complex on the target DNA. Next, a donor sequence of nucleic acid is introduced into the same stem cell. As a result, the guide RNA and Cas enzyme colocalize with the target DNA, Cas enzyme cleaves the target DNA, and the donor sequence of nucleic acid is embedded in the target DNA. Thus, altered DNA is obtained in the specified stem cell.

EFFECT: present invention allows for the use of specially structured nucleases in the construction of human induced pluripotent stem cells.

35 cl, 50 dwg, 9 tbl, 20 ex