Gene expression platform for synthetic biology in the human pathogen Streptococcus pneumoniae.
The human pathogen Streptococcus pneumoniae (pneumococcus) is a bacterium that owes its success to complex gene expression regulation patterns on both the cellular and the population level. Expression of virulence factors enables a mostly hazard-free presence of the commensal, in balance with the host and niche competitors. Under specific circumstances, changes in this expression can result in a more aggressive behavior and the reversion to the invasive form as pathogen. These triggering conditions are very difficult to study due to the fact that environmental cues are often unknown or barely possible to simulate outside the host (in vitro). An alternative way of investigating expression patterns is found in synthetic biology approaches of reconstructing regulatory networks that mimic an observed behavior with orthogonal components. Here, we created a genetic platform suitable for synthetic biology approaches in S. pneumoniae and characterized a set of standardized promoters and reporters. We show that our system allows for fast and easy cloning with the BglBrick system and that reliable and robust gene expression after integration into the S. pneumoniae genome is achieved. In addition, the cloning system was extended to allow for direct linker-based assembly of ribosome binding sites, peptide tags, and fusion proteins, and we called this new generally applicable standard "BglFusion". The gene expression platform and the methods described in this study pave the way for employing synthetic biology approaches in S. pneumoniae.
Molecular Genetics Group, Groningen Biomolecular Sciences and Biotechnology Institute, Centre for Synthetic Biology, University of Groningen, Nijenborgh 7, 9747 AG, Groningen, The Netherlands.
Robin A
RA
Sorg
Molecular Genetics Group, Groningen Biomolecular Sciences and Biotechnology Institute, Centre for Synthetic Biology, University of Groningen, Nijenborgh 7, 9747 AG, Groningen, The Netherlands.
Oscar P
OP
Kuipers
Molecular Genetics Group, Groningen Biomolecular Sciences and Biotechnology Institute, Centre for Synthetic Biology, University of Groningen, Nijenborgh 7, 9747 AG, Groningen, The Netherlands.
Jan-Willem
JW
Veening
D003001:Cloning, Molecular / Q000379:methods*; D015964:Gene Expression Regulation, Bacterial / Q000235:genetics*; D005822:Genetic Vectors / Q000235:genetics*; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins; D013296:Streptococcus pneumoniae / Q000235:genetics* / Q000378:metabolism; D058615:Synthetic Biology / Q000379:methods*
D011993:Recombinant Fusion Proteins
BglFusion; BioBrick; GFP; Golden Gate Shuffling; luciferase; mKate2