清脆的
生物
基因组编辑
基因组
双歧杆菌
计算生物学
遗传学
基因组工程
基因组
表观遗传学
基因组学
基因
细菌
乳酸菌
作者
Meichen Pan,Wesley Morovic,Claudio Hidalgo-Cantabrana,Avery Roberts,Kimberly K. O. Walden,Yong Meng Goh,Rodolphe Barrangou
出处
期刊:Proceedings of the National Academy of Sciences
日期:2022-07-20
卷期号:119 (30)
被引量:8
标识
DOI:10.1073/pnas.2205068119
摘要
Bifidobacterium is a commensal bacterial genus ubiquitous in the human gastrointestinal tract, which is associated with a range of health benefits. The advent of CRISPR-based genome editing technologies provides opportunities to investigate the genetics of important bacteria and transcend the lack of genetic tools in bifidobacteria to study the basis for their health-promoting attributes. Here, we repurpose the endogenous type I-G CRISPR-Cas system and adopt an exogenous CRISPR base editor for genome engineering in B. animalis subsp. lactis, demonstrating that both genomic and epigenetic contexts drive editing outcomes across strains. We reprogrammed the endogenous type I-G system to screen for naturally occurring large deletions up to 27 kb and to generate a 500-bp deletion in tetW to abolish tetracycline resistance. A CRISPR-cytosine base editor was optimized to install C•G-to-T•A amber mutations to resensitize multiple B. lactis strains to tetracycline. Remarkably, we uncovered epigenetic patterns that are distributed unevenly among B. lactis strains, despite their genomic homogeneity, that may contribute to editing efficiency variability. Insights were also expanded to Bifidobacterium longum subsp. infantis to emphasize the broad relevance of these findings. This study highlights the need to develop individualized CRISPR-based genome engineering approaches for distinct bacterial strains and opens avenues for engineering of next generation probiotics.
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