Development of Whole Genome‐Scale Base Editing Toolbox to Promote Efficiency of Extracellular Electron Transfer in Shewanella oneidensis MR‐1. Issue 3 (18th February 2022)

• Record Type: Journal Article
• Authors: Chen, Yaru; Fang, Lixia; Ying, Xiang; Cheng, Meijie; Wang, Lin; Sun, Panxing; Zhang, Zhaoyu; Shi, Liang; Cao, Yingxiu; Song, Hao
• Is Part Of: Advanced biology. Volume 6:Issue 3(2022)
• Page Start: n/a
• Page End: n/a
• Publication Date: 2022-02-18
• Subjects: biofilms -- cell morphology -- CRISPR base editing -- electroactive microorganisms -- extracellular electron transfer; Molecular biology -- Periodicals; Systems biology -- Periodicals; Biological systems -- Periodicals; Biotechnology -- Periodicals; Bioengineering -- Periodicals; Biomedical engineering -- Periodicals; 660.6
• DOI: 10.1002/adbi.202101296
• Languages: English
• ISSNs: 2701-0198

Abstract

Abstract: Shewanella oneidensis MR‐1, as a model electroactive microorganism (EAM) for extracellular electron transfer (EET) study, plays a key role in advancing practical applications of bio‐electrochemical systems (BES). Efficient genome‐level manipulation tools are vital to promote EET efficiency; thus, a powerful and rapid base editing toolbox in S. oneidensis MR‐1 is developed. Firstly a CRISPR/dCas9‐AID base editor that shows a relatively narrow editing window restricted to the "− 20 to − 16" range upstream of the protospacer adjacent motif (PAM) is constructed. Cas9 is also confined by its native PAM requirement, NGG. Then to expand the editable scope, the sgRNA and the Cas‐protein to broaden the editing window to "− 22 to − 9" upstream of the PAM are engineered, and the PAM field to NNN is opened up. Consequently, the coverage of the editable gene is expanded from 89% to nearly 100% in S. oneidensis MR‐1. This whole g enome‐scale cytidine deaminase‐based base editing toolbox (WGcBE) is applied to regulate the cell length and the biofilm morphology, which enhances the EET efficiency by 6.7‐fold. WGcBE enables an efficient deactivation of genes with full genome coverage, which would contribute to the in‐depth and multi‐faceted EET study in Shewanella . Abstract : Through systematic optimization of the editing window and the protospacer adjacent motif profiles of the base editor, this study expands the coverage of the editable scope to the whole genome of the model electroactive microorganism Shewanella oneidensis MR‐1. The strains with silenced lifespan‐ or biofilm‐related genes edited by the base editing toolbox show improved extracellular electron transfer efficiency than the wild‐type strain.