| Autor: |
Yulin Zhang, Yang Zheng, Qiwen Hu, Zhen Hu, Jiyuan Sun, Ping Cheng, Xiancai Rao, Xiao-Ran Jiang |
| Jazyk: |
angličtina |
| Rok vydání: |
2024 |
| Předmět: |
|
| Zdroj: |
Synthetic and Systems Biotechnology, Vol 9, Iss 3, Pp 586-593 (2024) |
| Druh dokumentu: |
article |
| ISSN: |
2405-805X |
| DOI: |
10.1016/j.synbio.2024.04.016 |
| Popis: |
Halomonas bluephagenesis TD serves as an exceptional chassis for next generation industrial biotechnology to produce various products. However, the simultaneous editing of multiple loci in H. bluephagenesis TD remains a significant challenge. Herein, we report the development of a multiple loci genome editing system, named CRISPR-deaminase-assisted base editor (CRISPR-BE) in H. bluephagenesis TD. This system comprises two components: a cytidine (CRISPR-cBE) and an adenosine (CRISPR-aBE) deaminase-based base editor. CRISPR-cBE can introduce a cytidine to thymidine mutation with an efficiency of up to 100 % within a 7-nt editing window in H. bluephagenesis TD. Similarly, CRISPR-aBE demonstrates an efficiency of up to 100 % in converting adenosine to guanosine mutation within a 7-nt editing window. CRISPR-cBE has been further validated and successfully employed for simultaneous multiplexed editing in H. bluephagenesis TD. Our findings reveal that CRISPR-cBE efficiently inactivated all six copies of the IS1086 gene simultaneously by introducing stop codon. This system achieved an editing efficiency of 100 % and 41.67 % in inactivating two genes and three genes, respectively. By substituting the Pcas promoter with the inducible promoter PMmp1, we optimized CRISPR-cBE system and ultimately achieved 100 % editing efficiency in inactivating three genes. In conclusion, our research offers a robust and efficient method for concurrently modifying multiple loci in H. bluephagenesis TD, opening up vast possibilities for industrial applications in the future. |
| Databáze: |
Directory of Open Access Journals |
| Externí odkaz: |
|
