Enhanced sleeping beauty transposons, kits and methods of transpositon

Abstract

The present invention relates to enhanced Sleeping Beauty-type transposons and methods of transposition. In particular the invention relates to a polynucleotide comprising a cargo nucleic acid flanked by a left and a right inverted repeat/direct repeat (IR/DR), wherein IR/DRs, having specific sequences, are recognized by a Sleeping Beauty transposase protein and the polynucleotide is capable of integrating into the DNA of a cell. The invention also relates to a kit for transposing a nucleic acid comprising said polynucleotide as well as to further components such as co-factors of transposition capable of depleting a component of the FACT (facilitates chromatin transcription) complex, namely, SSRP1 and/or SUPT16H/SPT16, or an inhibitor of cathepsin selected from the group comprising H,S,V, and L; or a cofactor capable of depleting or inhibiting HSP90; or a factor temporally arresting cells cell cycle in cell cycle phase G0/G1, G1/S, or G2/M; or a factor inhibiting the ubiquitination of PCNA, or cells wherein these components have been knocked down or inhibited,or the cell cyle arrested in any of said stages. Alternatively or additionally, the kit may comprise as a co-factor of transposition an agent capable of increasing concentration and/or signaling of ATR or a cell wherein concentrationand/or signaling of ATR are increased. The invention further provides methods using said transposon polynucleotide as well as host cells and pharmaceutical compositions. It also relates to use of said co-factors of transposition or specific cells for enhancing transposition efficiencies, e.g., for preparing genetically modified nucleic acids or cells.

Dependencies are inferred from the claim text.