Periodic Reporting for period 3 - BIONICbacteria (Integrating a novel layer of synthetic biology tools in Pseudomonas, inspired by bacterial viruses)
Okres sprawozdawczy: 2022-09-01 do 2024-02-29
We aim to i) exploit phage-encoded genetic circuits as synthetic biology biobricks and new biotechnological chassis; ii) build synthetic phage modulators as novel payloads to regulate bacterial metabolism in a targeted manner; and iii) integrate the new circuits to create designer bacteria for applications in industrial fermentations and vaccine design. This innovative approach will allow us to domesticate Pseudomonas strains with the goal of addressing key problems in industrial fermentations and vaccine development for society and industry.
• ONT-cappable-seq (Putzeys et al., in subm.) explores the primary prokaryotic transcriptional landscape by combining primary RNA enrichment with Nanopore long-read sequencing technology. This technology has been key to obtain the transcriptional blueprints envisioned in the WP1 and their application in WP2. We have also designed and made available the bioinformatics scripts (available on https://github.com/LoGT-KULeuven/ONT-cappable-seq) to analyze ONT-cappable-seq data.
• SAPPHIRE (Coppens and Lavigne, 2020 in BMC Bioinformatics) and its latest release SAPPHIRE.CNN (in prep.) are promoter prediction software specific for σ70 promoters in Pseudomonas and Salmonella. These tools outperform other promoter predictive software for the mentioned species. This technology has contributed to the identification of novel promoters and the confirmation of TSS identified by ONT-cappable-seq (WP1).
• SEVAtile is a standardized DNA assembly method optimized for Pseudomonas and compatible with the SEVA‐vector backbone (Lammens et al., 2021 in Microb. Biotechnol.). The implementation of this system in Pseudomonas enables the rapid and standardized assembly of genetic parts to create genetic circuits for analysis in non-model hosts.