Purified Ref polypeptides with increased nuclease site-specific targeting activity, recombinant nucleic acids and cells for expression of such Ref polypeptides, and methods for using the Ref polypeptides in combination with RecA protein and variants thereof to effect targeted nuclease cleavage of a DNA duplex are disclosed.

Purified Ref polypeptides with increased nuclease site-specific targeting activity, recombinant nucleic acids and cells for expression of such Ref polypeptides, and methods for using the Ref polypeptides in combination with RecA protein and variants thereof to effect targeted nuclease cleavage of a DNA duplex are disclosed.

Purified Ref polypeptides with increased nuclease site-specific targeting activity, recombinant nucleic acids and cells for expression of such Ref polypeptides, and methods for using the Ref polypeptides in combination with RecA protein and variants thereof to effect targeted nuclease cleavage of a DNA duplex are disclosed.

According to one embodiment, there is provided a method of producing a gene modification T cell (CAR-T cells) which expresses a chimeric antigen receptor (CAR). This method includes stimulating a cell population containing a T cell with an antibody which activates the T cell, bringing the cell population into contact with a nucleic acid-introducing carrier, wherein the nucleic acid-introducing carrier containing a lipid particle, a first nucleic acid and containing a CAR gene, and a second nucleic acid containing a transposase gene encapsulated in the lipid particle, and culturing the cell population after the contacting.