Method For The Selection Of Cells Based On Crispr/Cas-Controlled Integration Of A Detectable Tag To A Target Protein


Ontology type: sgo:Patent     


Patent Info

DATE

2020-07-09T00:00

AUTHORS

HAAS ALEXANDER

ABSTRACT

Herein is reported a method for providing cells that express a fusion protein from a marker protein and a cell-endogenous target protein, consisting of the steps a) transfecting the cells i) with a Cas9-encoding plasmid additionally containing a nucleic acid which is resistant to a first selection agent, ii) with a circular donor plasmid containing a first nucleic acid which confers resistance to a second selection reagent, and a second nucleic acid which encodes the marker protein and which is flanked 3' and 5' by nucleic acids homologous to the integration site in the cell, iii) a suitable synthetic crRNA, and iv) a suitable synthetic tracrRNA, b) cultivating the cells in the presence of the first and the second selection reagent, and c) selecting cells which under the conditions of step b) carry out cell division, and thereby providing/producing cells that express a fusion protein of a marker protein and a cell- endogenous target protein. More... »

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