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AssigneeMax-DelbrüCk-Centrum FüR Molekulare Medizin In Der Helmholtz-Gemeinschaft
InventorsWolfgang UCKERT, Mario BUNSE, Julian CLAUSS, Zsuzsanna IZSVÁK
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The present invention relates to the field of genetic engineering, in particular, to a transposon-based transfection kit suitable for transfection of primary cells, such as T cells, comprising mRNA encoding a transposase, or reagents for generating mRNA encoding said transposase, as well as minicircle DNA comprising the transposon. The invention also relates to a nucleic acid, preferably, a DNA minicircle, comprising a transposon, wherein the transposon encodes a protein and at least one miRNA, wherein the sequences encoding the miRNA are located in an intron and expression of the protein and the miRNA is regulated by the same promoter. The invention also provides a population of cells obtainable with the method of the invention. Methods of transfection are also provided, as well as medical use, e.g. in immunotherapy, in particular, in adoptive T cell therapy or T cell receptor (TCR) or chimeric antigen receptor (CAR) gene therapy.
A kit comprising
The kit of any of the preceding claims, wherein the transposase is selected from the group of class II transposable elements comprising piggyBac, Tol2 and MI/mariner-type transposons comprising Frog Prince and Sleeping Beauty transposase, preferably, Sleeping Beauty transposase, most preferably, SB100X.
A nucleic acid comprising a transposon, wherein the transposon encodes a protein and a miRNA, wherein the nucleic acid encoding the protein comprises an intron comprising sequences encoding the miRNA, wherein expression of the protein and the miRNA is regulated by the same promoter, wherein the nucleic acid is selected from the group comprising a plasmid or minicircle DNA.
Dependencies are inferred from the claim text.