DNA Double-Strand Break-Induced Gene Amplification in Yeast
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DNA Double-Strand Break-Induced Gene Amplification in Yeast. / Strucko, Tomas; Lisby, Michael; Mortensen, Uffe Hasbro.
Homologous Recombination: Methods and Protocols. red. / Andrés Aguilera; Aura Carreira. Humana Press, 2021. s. 239-252 (Methods in Molecular Biology, Bind 2153).Publikation: Bidrag til bog/antologi/rapport › Bidrag til bog/antologi › Forskning › fagfællebedømt
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TY - CHAP
T1 - DNA Double-Strand Break-Induced Gene Amplification in Yeast
AU - Strucko, Tomas
AU - Lisby, Michael
AU - Mortensen, Uffe Hasbro
PY - 2021
Y1 - 2021
N2 - Precise control of the gene copy number in the model yeast Saccharomyces cerevisiae may facilitate elucidation of enzyme functions or, in cell factory design, can be used to optimize production of proteins and metabolites. Currently, available methods can provide high gene-expression levels but fail to achieve accurate gene dosage. Moreover, strains generated using these methods often suffer from genetic instability resulting in loss of gene copies during prolonged cultivation. Here we present a method, CASCADE, which enables construction of strains with defined gene copy number. With our present system, gene(s) of interest can be amplified up to nine copies, but the upper copy limit of the system can be expanded. Importantly, the resulting strains can be stably propagated in selection-free media.
AB - Precise control of the gene copy number in the model yeast Saccharomyces cerevisiae may facilitate elucidation of enzyme functions or, in cell factory design, can be used to optimize production of proteins and metabolites. Currently, available methods can provide high gene-expression levels but fail to achieve accurate gene dosage. Moreover, strains generated using these methods often suffer from genetic instability resulting in loss of gene copies during prolonged cultivation. Here we present a method, CASCADE, which enables construction of strains with defined gene copy number. With our present system, gene(s) of interest can be amplified up to nine copies, but the upper copy limit of the system can be expanded. Importantly, the resulting strains can be stably propagated in selection-free media.
KW - CEN.PK
KW - DNA double-strand break
KW - Gene amplification
KW - Gene targeting
KW - Homology-directed recombination
KW - I-SceI nuclease
KW - Metabolic engineering
KW - Saccharomyces cerevisiae
U2 - 10.1007/978-1-0716-0644-5_17
DO - 10.1007/978-1-0716-0644-5_17
M3 - Book chapter
C2 - 32840784
AN - SCOPUS:85089925285
SN - 978-1-0716-0643-8
T3 - Methods in Molecular Biology
SP - 239
EP - 252
BT - Homologous Recombination
A2 - Aguilera, Andrés
A2 - Carreira, Aura
PB - Humana Press
ER -
ID: 249857300