关键词: 短小芽孢杆菌, 顺式作用元件, 转录组, UgRNA/Cas9多基因编辑, 代谢组

Abstract:

【Objective】 Due to the negative impact of long-term synthetic fertilizer application on the agricultural functions of rhizosphere microorganisms, we aim to develop an effective approach to enhance these functions in a wild-type Bacillus pumilus isolated from plant roots.【Method】 A novel CRISPR/Cas multi-gene editing system, UgRNA/Cas9, was used to disrupt the genome-scale cis-acting catabolite-responsive element（CRE）, aiming to alter the carbon source selectivity of B. pumilus. 【Result】 DNA sequencing results revealed that CRE-like sites in the partial genes of the carbon and secondary metabolic pathways underwent mutations including deletions, insertions, transitions, and transversions. Comparative metabolomic and transcriptomic analyses suggest a potential for the biosynthesis of pigments, surfactin, and bacilysin through the pentose phosphate and amino acid pathways. The introduction of UgRNA/Cas9-mediated edits into the CREs enhanced the ability of the strain to better adhere to the root, thereby promoting plant growth and strengthening resistance to pathogens. 【Conclusion】 Mutations in the genome-wide CRE sequence of LG3145 altered the direction of carbon metabolism flow in the bacterium, resulting in the establishment of a mutually beneficial relationship between the bacterium and plants.

Key words: Bacillus pumilus, cis-acting elements, transcriptome, UgRNA/Cas9 multi-gene editing, metabolome