An Introduced RNA-Only Approach for Plasmid Curing via the CRISPR-Cpf1 System in <i>Saccharomyces cerevisiae</i>
The CRISPR-Cas system has been widely used for genome editing due to its convenience, simplicity and flexibility. Using a plasmid-carrying Cas protein and crRNA or sgRNA expression cassettes is an efficient strategy in the CRISPR-Cas genome editing system. However, the plasmid remains in the cells a...
| Päätekijät: | , , |
|---|---|
| Aineistotyyppi: | Artikkeli |
| Kieli: | English |
| Julkaistu: |
MDPI AG
2023-10-01
|
| Sarja: | Biomolecules |
| Aiheet: | |
| Linkit: | https://www.mdpi.com/2218-273X/13/10/1561 |
| _version_ | 1827721516645613568 |
|---|---|
| author | Bo-Chou Chen Yu-Zhen Chen Huan-Yu Lin |
| author_facet | Bo-Chou Chen Yu-Zhen Chen Huan-Yu Lin |
| author_sort | Bo-Chou Chen |
| collection | DOAJ |
| description | The CRISPR-Cas system has been widely used for genome editing due to its convenience, simplicity and flexibility. Using a plasmid-carrying Cas protein and crRNA or sgRNA expression cassettes is an efficient strategy in the CRISPR-Cas genome editing system. However, the plasmid remains in the cells after genome editing. Development of general plasmid-curing strategies is necessary. Based on our previous CRISPR-Cpf1 genome-editing system in <i>Saccharomyces cerevisiae</i>, the crRNA, designed for the replication origin of the CRISPR-Cpf1 plasmid, and the ssDNA, as a template for homologous recombination, were introduced for plasmid curing. The efficiency of the plasmid curing was 96 ± 4%. In addition, we further simplified the plasmid curing system by transforming only one crRNA into <i>S. cerevisiae</i>, and the curing efficiency was about 70%. In summary, we have developed a CRISPR-mediated plasmid-curing system. The RNA-only plasmid curing system is fast and easy. This plasmid curing strategy can be applied in broad hosts by designing crRNA specific for the replication origin of the plasmid. The plasmid curing system via CRISPR-Cas editing technology can be applied to produce traceless products without foreign genes and to perform iterative processes in multiple rounds of genome editing. |
| first_indexed | 2024-03-10T21:23:32Z |
| format | Article |
| id | doaj.art-6bf76622499a4c06aac551af47c17147 |
| institution | Directory Open Access Journal |
| issn | 2218-273X |
| language | English |
| last_indexed | 2024-03-10T21:23:32Z |
| publishDate | 2023-10-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Biomolecules |
| spelling | doaj.art-6bf76622499a4c06aac551af47c171472023-11-19T15:51:00ZengMDPI AGBiomolecules2218-273X2023-10-011310156110.3390/biom13101561An Introduced RNA-Only Approach for Plasmid Curing via the CRISPR-Cpf1 System in <i>Saccharomyces cerevisiae</i>Bo-Chou Chen0Yu-Zhen Chen1Huan-Yu Lin2Bioresource Collection and Research Center, Food Industry Research and Development Institute, Hsinchu 300, TaiwanDepartment of Food Science and Technology, Hungkuang University, Taichung 433, TaiwanBioresource Collection and Research Center, Food Industry Research and Development Institute, Hsinchu 300, TaiwanThe CRISPR-Cas system has been widely used for genome editing due to its convenience, simplicity and flexibility. Using a plasmid-carrying Cas protein and crRNA or sgRNA expression cassettes is an efficient strategy in the CRISPR-Cas genome editing system. However, the plasmid remains in the cells after genome editing. Development of general plasmid-curing strategies is necessary. Based on our previous CRISPR-Cpf1 genome-editing system in <i>Saccharomyces cerevisiae</i>, the crRNA, designed for the replication origin of the CRISPR-Cpf1 plasmid, and the ssDNA, as a template for homologous recombination, were introduced for plasmid curing. The efficiency of the plasmid curing was 96 ± 4%. In addition, we further simplified the plasmid curing system by transforming only one crRNA into <i>S. cerevisiae</i>, and the curing efficiency was about 70%. In summary, we have developed a CRISPR-mediated plasmid-curing system. The RNA-only plasmid curing system is fast and easy. This plasmid curing strategy can be applied in broad hosts by designing crRNA specific for the replication origin of the plasmid. The plasmid curing system via CRISPR-Cas editing technology can be applied to produce traceless products without foreign genes and to perform iterative processes in multiple rounds of genome editing.https://www.mdpi.com/2218-273X/13/10/1561CRISPR-Cpf1plasmid-curinggenome-editing<i>Saccharomyces cerevisiae</i> |
| spellingShingle | Bo-Chou Chen Yu-Zhen Chen Huan-Yu Lin An Introduced RNA-Only Approach for Plasmid Curing via the CRISPR-Cpf1 System in <i>Saccharomyces cerevisiae</i> Biomolecules CRISPR-Cpf1 plasmid-curing genome-editing <i>Saccharomyces cerevisiae</i> |
| title | An Introduced RNA-Only Approach for Plasmid Curing via the CRISPR-Cpf1 System in <i>Saccharomyces cerevisiae</i> |
| title_full | An Introduced RNA-Only Approach for Plasmid Curing via the CRISPR-Cpf1 System in <i>Saccharomyces cerevisiae</i> |
| title_fullStr | An Introduced RNA-Only Approach for Plasmid Curing via the CRISPR-Cpf1 System in <i>Saccharomyces cerevisiae</i> |
| title_full_unstemmed | An Introduced RNA-Only Approach for Plasmid Curing via the CRISPR-Cpf1 System in <i>Saccharomyces cerevisiae</i> |
| title_short | An Introduced RNA-Only Approach for Plasmid Curing via the CRISPR-Cpf1 System in <i>Saccharomyces cerevisiae</i> |
| title_sort | introduced rna only approach for plasmid curing via the crispr cpf1 system in i saccharomyces cerevisiae i |
| topic | CRISPR-Cpf1 plasmid-curing genome-editing <i>Saccharomyces cerevisiae</i> |
| url | https://www.mdpi.com/2218-273X/13/10/1561 |
| work_keys_str_mv | AT bochouchen anintroducedrnaonlyapproachforplasmidcuringviathecrisprcpf1systeminisaccharomycescerevisiaei AT yuzhenchen anintroducedrnaonlyapproachforplasmidcuringviathecrisprcpf1systeminisaccharomycescerevisiaei AT huanyulin anintroducedrnaonlyapproachforplasmidcuringviathecrisprcpf1systeminisaccharomycescerevisiaei AT bochouchen introducedrnaonlyapproachforplasmidcuringviathecrisprcpf1systeminisaccharomycescerevisiaei AT yuzhenchen introducedrnaonlyapproachforplasmidcuringviathecrisprcpf1systeminisaccharomycescerevisiaei AT huanyulin introducedrnaonlyapproachforplasmidcuringviathecrisprcpf1systeminisaccharomycescerevisiaei |