C - Chemistry – Metallurgy – 12 – N
Patent
C - Chemistry, Metallurgy
12
N
C12N 15/63 (2006.01) C12N 15/10 (2006.01) C12N 15/67 (2006.01) C12N 15/90 (2006.01) C12P 19/34 (2006.01) C12P 21/02 (2006.01)
Patent
CA 2152371
A method of amplifying in vivo a DNA sequence B present in a genome of a parent cell, comprising (a) integrating in a genome of said cell a DNA construct comprising the structure C-M-A-D, in which both A and C denote a DNA sequence which is homologous with a genomic DNA fragment either flanking or overlapping the DNA sequence B to be amplified or being a subsequence of the DNA sequence B constituting one of the ends of said sequence B, the sequence C being located in the opposite end of the sequence B as compared to A, D denotes a DNA sequence which is homologous with a genomic DNA fragment located distal for C as compared to B, and M denotes a DNA sequence encoding a selection marker, (b) selecting for cells in wich the DNA sequence M has been integrated in the genome, which cells comprise, in any orientation, the structure A-B-C-M-A-D, and (c) propagating the cells selected in step (b) under increasing selection pressure to obtain a cell which has obtained an increased number of genomically integrated copies of the DNA sequences B and M.
Mccarthy Tetrault Llp
Novo Nordisk A/s
Novozymes A/s
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