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US 2020/0263188 A1Published application
Methods for genetic engineering kluyveromyces host cells
AssigneeAmyris, Inc.
InventorYoseph TSEGAYE
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Abstract
The invention provides methods for genetically engineering Kluyveromyces. The methods can be used to genetically engineer Kluyveromyces to produce and secrete full-length antibodies or antibody fragments. The invention also provides methods for production and secretion of antibodies.
Claims25
A method for modifying a target site in a Kluyveromyces host cell genome, the method comprising:
- (a) contacting a Kluyveromyces host cell, which has a reduced non-homologous end joining (NHEJ) activity with:
- (i) a nuclease capable of cleaving the target site;
- and
- (ii) a donor DNA molecule comprising a nucleic acid sequence encoding a polypeptide, wherein the donor DNA molecule is capable of homologous recombination at the cleaved target site, whereby homologous recombination in the host cell results in integration of the donor linear nucleic acid at the target site;
- and
- (b) selecting a transformed host cell in which the donor DNA molecule is integrated into the target site and the host cell secretes the polypeptide encoded by the nucleic acid sequence.
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Dependencies are inferred from the claim text.