Recycling selectable markers via Cre/loxP system for constructing Komagataella phaffii strains co-expressing multiple proteins
© 2024. The Author(s), under exclusive licence to Springer Nature B.V..
OBJECTIVE: A convenient strategy was developed to recycle selectable markers using Cre/loxP system for constructing Komagataella phaffii strains co-expressing multiple proteins.
RESULTS: A plasmid in this strategy was generated from pPICZαA with integration of lox71-Sh ble-lox66. Firstly, the plasmid was inserted with one target protein gene and then transformed into K. phaffii KM71. Secondly, the auxiliary plasmid pPICZαA/cre/his4 containing CRE recombinase gene was further chromosomally inserted to Sh ble gene therein. Finally, methanol induction was conducted to produce CRE for Cre/loxP-mediated recombination, and consequently, the sequence between lox71 and lox66 was deleted, leading to recycling of ZeoR and His- markers. Then the resulted strain expressing the one target protein was used as the host to which another target protein gene could be inserted by the same procedures.
CONCLUSIONS: With easy manipulation, the method was effective in recycling of the selectable markers, and consequently two protein genes were sequential integrated chromosomally and successfully co-expressed in the yeast.
| Medienart: |
E-Artikel |
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| Erscheinungsjahr: |
2024 |
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| Erschienen: |
2024 |
| Enthalten in: |
Zur Gesamtaufnahme - volume:46 |
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| Enthalten in: |
Biotechnology letters - 46(2024), 3 vom: 28. Apr., Seite 399-407 |
| Sprache: |
Englisch |
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| Beteiligte Personen: |
Wang, Weixian [VerfasserIn] |
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| Links: |
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| Themen: |
Co-express multi-protein |
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| Anmerkungen: |
Date Completed 27.04.2024 Date Revised 27.04.2024 published: Print-Electronic Citation Status MEDLINE |
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| doi: |
10.1007/s10529-024-03466-3 |
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| funding: |
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| Förderinstitution / Projekttitel: |
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| 245 | 1 | 0 | |a Recycling selectable markers via Cre/loxP system for constructing Komagataella phaffii strains co-expressing multiple proteins |
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| 500 | |a published: Print-Electronic | ||
| 500 | |a Citation Status MEDLINE | ||
| 520 | |a © 2024. The Author(s), under exclusive licence to Springer Nature B.V. | ||
| 520 | |a OBJECTIVE: A convenient strategy was developed to recycle selectable markers using Cre/loxP system for constructing Komagataella phaffii strains co-expressing multiple proteins | ||
| 520 | |a RESULTS: A plasmid in this strategy was generated from pPICZαA with integration of lox71-Sh ble-lox66. Firstly, the plasmid was inserted with one target protein gene and then transformed into K. phaffii KM71. Secondly, the auxiliary plasmid pPICZαA/cre/his4 containing CRE recombinase gene was further chromosomally inserted to Sh ble gene therein. Finally, methanol induction was conducted to produce CRE for Cre/loxP-mediated recombination, and consequently, the sequence between lox71 and lox66 was deleted, leading to recycling of ZeoR and His- markers. Then the resulted strain expressing the one target protein was used as the host to which another target protein gene could be inserted by the same procedures | ||
| 520 | |a CONCLUSIONS: With easy manipulation, the method was effective in recycling of the selectable markers, and consequently two protein genes were sequential integrated chromosomally and successfully co-expressed in the yeast | ||
| 650 | 4 | |a Journal Article | |
| 650 | 4 | |a Komagataella phaffii | |
| 650 | 4 | |a Pichia pastoris | |
| 650 | 4 | |a Co-express multi-protein | |
| 650 | 4 | |a Cre/loxP | |
| 650 | 4 | |a Selectable marker | |
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| 700 | 1 | |a Han, Minghai |e verfasserin |4 aut | |
| 700 | 1 | |a Zhu, Guofei |e verfasserin |4 aut | |
| 700 | 1 | |a Liu, Xiaohui |e verfasserin |4 aut | |
| 700 | 1 | |a Zhao, Tianming |e verfasserin |4 aut | |
| 700 | 1 | |a Ma, Xiaoyan |e verfasserin |4 aut | |
| 700 | 1 | |a Gong, Xun |e verfasserin |4 aut | |
| 700 | 1 | |a Xu, Cunbin |e verfasserin |4 aut | |
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