Thanks! First challenge is to expand the substrate acceptance range in the cell factory

🔬 Start: Jan 2026 | 1-year funded
🏭 Work with AMBR + 10–30L reactors
📈 TRY optimization | in vivo polymerization | TEA & LCA
💼 Great stepping stone to fellowships (NNF, EU, etc.)

LinkedIn post with more info: www.linkedin.com/posts/enrico...

We had great support from our labs in developing this toolkit. Thanks to @pabnik.bsky.social, @nicoc-micsynmet.bsky.social, Sjoerd, Luc, Angela, Chase, Harrison, Raymond, John, and Wei!

The plasmids will be available on Addgene. Feel free to reach out in case you want to try them!

These results demonstrates that a new state-of-the-art is available for making gene deletions in this promising bug.
With a turnaround time that is 50% shorter than the current systems available, we believe that SIBR-Cas9 and SIBR2.0-Cas12a will help using C. necator in new P2X applications! 8/n

SIBR2.0 was then functional and we moved testing it for Cas12a. The new architectures revealed to be tight, allowing precise and controllable induction of cas12a as well, with similar efficiencies to the ones observed for Cas9! 7/n

Here is where SIBR2.0 comes into place. Simona and Costas developed a script for strategically moving the intron position within the CDS to get rid of such hidden translation initiation starts. This was tested first in E. coli using GFP as readout. 6/n

Then, we moved at testing if SIBR would work also on Cas12a. To our surprise, induction during the targeting assay revealed a leaky cas12a expression despite the presence of the intron with the STOP codon. This was because of a hidden translation start site that required some adjustments. 5/n

We started by testing the SIBR setup on Cas9. This was because we already had evidence that it worked before, although poorly www.sciencedirect.com/science/arti...
Targeting worked fine & by combining SIBR w/ Cas9, we achieved high editing efficiencies of >80% in two loci! Great start! 4/n

Teaming up with Simona and Costas, we decided to apply the SIBR technology that Costas developed during his PhD to "pause" Cas counter-selection and allow endogenous homologous recombination (HR) to happen first. Our suspect was that C. necator is also poor at HR academic.oup.com/nar/article/... 3/n

C. necator holds potential for the conversion and valorization of CO2 for its ability to grow on this substrate using power-to-X feedstocks (like hydrogen or formate). However, making gene deletions in this organism has always been a pain due to its limited gene deletion toolkit 2/n