Genetic toolsets for non-model, phylogenetically diverse gut Clostridia enable precise manipulation of microbiota gene expression

Gut Clostridia are abundant members of the human microbiota but remain difficult to genetically manipulate due to limited molecular tools. To enable functional studies across phylogenetically diverse gut Clostridial species, this study aimed to identify strong promoters capable of driving robust gene expression. We established a NanoLuc luciferase–based screening platform to experimentally test promoter activity. Prior to screening, candidate promoters were computationally prioritized using meta-genomic and transcriptomic datasets, under the assumption that promoters associated with highly expressed genes are more likely to be strong and broadly active. This R code implements the promoter prioritization pipeline, ranking candidate promoters based on expression strength and consistency across datasets.