A MATQ-seq-Based Protocol for Single-Cell RNA-seq in Bacteria.

Area of Science:

• Microbiology
• Genomics
• Molecular Biology

Background:

• Microbial adaptation relies on phenotypic heterogeneity, but bulk gene studies mask individual cell differences.
• Current methods average gene expression, limiting understanding of microbial responses.
• Single-cell resolution is crucial for studying microbial diversity and adaptation.

Purpose of the Study:

• To develop and validate a bacterial single-cell RNA sequencing (scRNA-seq) workflow.
• To adapt the MATQ-seq protocol for microbial applications.
• To enable unbiased, whole-transcriptome analysis at the single-cell level for bacteria.

Main Methods:

• Adaptation of the multiple annealing and dC-tailing-based quantitative single-cell RNA sequencing (MATQ-seq) protocol for bacteria.
• Application of the workflow to human pathogens: Salmonella enterica serovar Typhimurium and Pseudomonas aeruginosa.
• Detailed protocol development for bacterial scRNA-seq.

Main Results:

• Successful implementation of a modified MATQ-seq workflow for bacterial scRNA-seq.
• Demonstration of the protocol's applicability to key human pathogens.
• Establishment of a method for unbiased, single-cell transcriptome analysis in bacteria.

Conclusions:

• Bacterial scRNA-seq protocols, like the one presented, are essential for dissecting phenotypic heterogeneity.
• This workflow provides a powerful tool for exploring microbial transcriptomes in diverse conditions.
• Opens new avenues for studying microbial adaptation in infection, persistence, ecology, and biofilms.