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Updated: Jul 11, 2025

RIBO-seq in Bacteria: a Sample Collection and Library Preparation Protocol for NGS Sequencing
Published on: August 7, 2021
A smart RBS library and its prediction model for robust and accurate fine-tuning of gene expression in Bacillus
Xiaolan Rao1, Dian Li1, Zhaowei Su1
1State Key Laboratory of Biocatalysis and Enzyme Engineering, Environmental Microbial Technology Center of Hubei Province, Hubei University, Wuhan 430062, PR China.
Abstract:
Bacillus species, such as Bacillus subtilis and Bacillus licheniformis, are important industrial bacteria. However, there is a lack of standardized and predictable genetic tools for convenient and reproducible assembly of genetic modules in Bacillus species to realize their full potential. In this study, we constructed a Ribosome Binding Site (RBS) library in B. licheniformis, which provides incremental regulation of expression levels over a 104-fold range. Additionally, we developed a model to quantify the resulting translation rates. We successfully demonstrated the robust expression of various target genes using the RBS library and showed that the model accurately predicts the translation rates of arbitrary coding genes. Importantly, we also extended the use of the RBS library and prediction model to B. subtilis, B. thuringiensis, and B. amyloliquefacie. The versatility of the RBS library and its prediction model enables quantification of biological behavior, facilitating reliable forward engineering of gene expression.
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