| Autor: |
Junyi Wang, Jingya Song, Cong Fan, Jiahao Duan, Kaiyuan He, Jifeng Yuan |
| Jazyk: |
angličtina |
| Rok vydání: |
2024 |
| Předmět: |
|
| Zdroj: |
BioDesign Research, Vol 6 (2024) |
| Druh dokumentu: |
article |
| ISSN: |
2693-1257 |
| DOI: |
10.34133/bdr.0030 |
| Popis: |
In the biotechnological industry, multicopy gene integration represents an effective strategy to maintain a high-level production of recombinant proteins and to assemble multigene biochemical pathways. In this study, we developed copper-induced in vivo gene amplification in budding yeast for multicopy gene expressions. To make copper as an effective selection pressure, we first constructed a copper-sensitive yeast strain by deleting the CUP1 gene encoding a small metallothionein-like protein for copper resistance. Subsequently, the reporter gene fused with a proline–glutamate–serine–threonine-destabilized CUP1 was integrated at the δ sites of retrotransposon (Ty) elements to counter the copper toxicity at 100 μM Cu2+. We further demonstrated the feasibility of modulating chromosomal rearrangements for increased protein expression under higher copper concentrations. In addition, we also demonstrated a simplified design of integrating the expression cassette at the CUP1 locus to achieve tandem duplication under high concentrations of copper. Taken together, we envision that this method of copper-induced in vivo gene amplification would serve as a robust and useful method for protein overproduction and metabolic engineering applications in budding yeast. |
| Databáze: |
Directory of Open Access Journals |
| Externí odkaz: |
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