Pathway integration and expression in host cells

US11674145B2 · kind B2 · utility

Assignee

• The Regents of the University of California · US

Inventors

• Yasuo Yoshikuni · Albany, US
• Gaoyan WANG · Pleasanton, US
• Zhiying Zhao · Danville, US
• Robert Cameron COATES · Lafayette, US
• Jan-Fang CHENG · Orinda, US
• David Robinson · New York, US

Key dates

• Filing date: Jan 24, 2017
• Grant date: Jun 13, 2023
• Expiry date: Jan 18, 2040

Classification

• Technology area (CPC C): Chemistry; Metallurgy
• CPC primary: C12R2001/425
• WIPO field: Biotechnology
• WIPO sector: Chemistry

Abstract

Provided herein are methods for integrating a gene of interest into a chromosome of a host cell. In some embodiments, the methods include introducing into a host cell a first plasmid comprising a transposase coding sequence and a donor sequence, which includes a selectable marker coding sequence flanked by a first and a second lox site and is itself flanked by inverted repeats recognized by the transposase. Following transposase-mediated chromosomal integration of the donor sequence into the host cell, a second plasmid is introduced, which comprises the gene of interest and a second selectable marker coding sequence, both flanked by a first and a second lox site. The gene of interest is chromosomally integrated into the host cell by recombinase-mediated cassette exchange (RMCE) between the donor sequence and the second plasmid via Cre-/cuc recombination. Further provided herein are host cells, vectors, and methods of producing a product related thereto.