Back to Search
Start Over
High-Throughput Transcriptional Characterization of Regulatory Sequences from Bacterial Biosynthetic Gene Clusters
- Source :
- ACS Synthetic Biology. 10:1859-1873
- Publication Year :
- 2021
- Publisher :
- American Chemical Society (ACS), 2021.
-
Abstract
- Recent efforts to sequence, survey, and functionally characterize the diverse biosynthetic capabilities of bacteria have identified numerous Biosynthetic Gene Clusters (BGCs). Genes found within BGCs are typically transcriptionally silent, suggesting their expression is tightly regulated. To better elucidate the underlying mechanisms and principles that govern BGC regulation on a DNA sequence level, we employed high-throughput DNA synthesis and multiplexed reporter assays to build and to characterize a library of BGC-derived regulatory sequences. Regulatory sequence transcription levels were measured in the Actinobacteria Streptomyces albidoflavus J1074, a popular model strain from a genus rich in BGC diversity. Transcriptional activities varied over 1000-fold in range and were used to identify key features associated with expression, including GC content, transcription start sites, and sequence motifs. Furthermore, we demonstrated that transcription levels could be modulated through coexpression of global regulatory proteins. Lastly, we developed and optimized a S. albidoflavus cell-free expression system for rapid characterization of regulatory sequences. This work helps to elucidate the regulatory landscape of BGCs and provides a diverse library of characterized regulatory sequences for rational engineering and activation of cryptic BGCs.
- Subjects :
- DNA, Bacterial
Transcription, Genetic
biology
Biomedical Engineering
General Medicine
Computational biology
Regulatory Sequences, Nucleic Acid
biology.organism_classification
Mycobacterium bovis
Biochemistry, Genetics and Molecular Biology (miscellaneous)
Streptomyces
Synthetic biology
Bacterial Proteins
Regulatory sequence
Transcription (biology)
Nucleotide Motifs
Sequence motif
Gene
GC-content
Gene Library
Sequence (medicine)
Subjects
Details
- ISSN :
- 21615063
- Volume :
- 10
- Database :
- OpenAIRE
- Journal :
- ACS Synthetic Biology
- Accession number :
- edsair.doi.dedup.....1491d5f47f69a54596c54e96047410c8
- Full Text :
- https://doi.org/10.1021/acssynbio.0c00639