An important aim of the iimena project is to develop methods that enable us to more efficiently get access to the potential new antibiotics produced by actinomycetes.
One of the most important developments in the recent years have been molecular tools using CRISPR technologies. iimena scientists have been on the forefront of developing such tools and sharing them with the international research community.
We are therefore very happy that a protocol now has been published in Nature Protocols. In the publication, titled “CRISPR–Cas9, CRISPRi and CRISPR-BEST-mediated genetic manipulation in streptomycetes”, users of the toolkit find very detailed step-by-step guides on how to most efficiently use our toolkit.
Link to interactive PDF of the protocol (No Nature Protocols subscription required):
Link to our blog post on the Nature Protocols and Methods Community:
The BEST is coming: Streptomyces natural products meet CRISPR toolkit
Links to the original research work the protocols are based and other CRISPR related papers of our group:
- Blin, K., Shaw, S., Tong, Y. & Weber, T. Designing sgRNAs for CRISPR-BEST base editing applications with CRISPy-web 2.0. Synth. Syst. Biotechnol. 5, 99-102, https://doi.org/10.1016/j.synbio.2020.05.005 (2020).
- Tong, Y., Weber, T. & Lee, S. Y. CRISPR/Cas-based genome engineering in natural product discovery. Nat. Prod. Rep. 36, 1262-1280, https://doi.org/10.1039/c8np00089a (2019).
- Tong, Y. et al. Highly efficient DSB-free base editing for streptomycetes with CRISPR-BEST. Proc. Natl. Acad. Sci. U. S. A. 116, 20366-20375, https://doi.org/10.1073/pnas.1913493116 (2019).
- Tong, Y., Lunde Robertsen, H., Blin, K., Weber, T. & Lee, S. Y. in Synthetic Metabolic Pathways: Methods and Protocols Vol. 1671 Methods in Molecular Biology (eds Michael Krogh Jensen & Jay Keasling) Ch. 11, 163-184 (Springer Science+Business Media LLC, 2018). https://doi.org/10.1007/978-1-4939-7295-1_11
- Blin, K., Pedersen, L. E., Weber, T. & Lee, S. Y. CRISPy-web: An online resource to design sgRNAs for CRISPR applications. Synth. Syst. Biotechnol. 1, 118-121, https://doi.org/10.1016/j.synbio.2016.01.003 (2016).
- Tong, Y., Charusanti, P., Zhang, L., Weber, T. & Lee, S. Y. CRISPR-Cas9 based engineering of actinomycetal genomes. ACS Synth. Biol. 4, 1020-1029, https://doi.org/10.1021/acssynbio.5b00038 (2015).