Methods for producing single insect cell clones
AssigneeuniQure biopharma B.V.
InventorsFiona Amanda Lucie MORENO, Anggakusuma, Hugo Federico CUETO ROJAS
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Abstract
The present invention relates to a method for producing a single cell clone of an insect cell comprising integrated into the genome of the cell at least one expression cassette for inducible expression of parvoviral Rep 78 and 52 proteins. The insect cells can be used for the production of parvoviral vectors by transfection constructs comprising parvoviral Cap and ITR sequences. Single cell clones obtained in the methods of the invention can be expanded for into a cell bank for the production of clinical grade parvoviral gene therapy vectors.
Claims22
A method for producing a single cell clone of an insect cell comprising at least one expression cassette for inducible expression of parvoviral Rep 78 and 52 proteins integrated into the genome of the cell, the method comprising:
- (a) preparing a series of sequential dilutions from a culture comprising a plurality of the insect cell;
- (b) seeding an aliquot from a dilution obtained in (a) into at least one culture container at a cell density of less than one insect cell per container;
- (c) incubating the culture container under conditions that are conducive to growth of the insect cell and that do not induce expression of the parvoviral Rep 78 and 52 proteins;
- and,
- (d) selecting at least one single cell clone from at least one culture container in which the insect cells have grown.
Dependencies are inferred from the claim text.