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Updated: Aug 31, 2025
Transcription Start Site Mapping Using Super-low Input Carrier-CAGE
Published on: June 26, 2019
Genome-wide transcription start site mapping in the facultative intracellular pathogen Brucella melitensis by
Fang Liu1, Yong-Fang Huang2, Chang-Xian Wu2
1Hongshan Laboratory, State Key Laboratory of Agricultural Microbiology, and Key laboratory of Preventive Veterinary Medicine in Hubei Province, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan 430070, China; CAS Key Laboratory of Molecular Virology and Immunology, Institut Pasteur of Shanghai, Chinese Academy of Sciences, Shanghai, China.
We developed Capping-seq to map transcription start sites (TSSs) in Brucella melitensis, identifying 2,369 TSSs. This advances understanding of gene regulation and brucellosis pathogenesis.
Area of Science:
- Microbiology
- Genomics
- Molecular Biology
Background:
- Brucella melitensis causes global zoonotic diseases (brucellosis), with intracellular survival hindering treatment.
- Limited gene annotation impedes research into bacterial gene expression regulation.
- Understanding transcription start sites (TSSs) is crucial for deciphering gene expression control.
Purpose of the Study:
- To develop and apply a novel genome-wide method for precise TSS mapping in B. melitensis.
- To improve gene annotation and explore potential functional regulatory sites.
- To gain systematic insight into the primary transcriptome of B. melitensis.
Main Methods:
- Developed Capping-seq, a novel genome-wide technique combining capped primary transcript capture with Single Molecule Real-Time (SMRT) sequencing.
- Applied Capping-seq to map transcription start sites (TSSs) in Brucella melitensis.
- Analyzed promoter elements (-35 and -10 regions) and 5' end features of identified transcripts.
Main Results:
- Identified 2,369 TSSs at single nucleotide resolution in B. melitensis.
- Revealed promoter consensus sequences (TTGNNN for -35, TATNNN for -10) and a purine preference at TSS positions.
- Found that 57% of genes have multiple TSSs, 47% possess long leader regions, and identified 52 leaderless metabolic genes, suggesting complex 5' end regulation.
Conclusions:
- Capping-seq is an effective technology for TSS determination in prokaryotes, including B. melitensis.
- The study provides a comprehensive characterization of the B. melitensis primary transcriptome.
- Findings establish a critical foundation for future research into Brucella gene regulation and pathogenesis.
