Team:Edinburgh OG/Parts
Parts
This year the iGEM Edinburgh_OG team focused on developing a modular toolkit using CRISPR systems and phages to re-sensitise antibiotic-resistant bacteria. As a BioBrick we submit the E. coli codon-optimised Staphylococcus aureus Cas9.
http://www.nature.com/news/genome-editing-revolution-my-whirlwind-year-with-crispr-1.19063
How does this part work?
- Our SaCas9 can be programmed to cleave specific target sequence followed by the PAM sequence (5’-NNGRRT-3’).
- To express SaCas9, it requires suitable machinery such as promoter, RBS,and terminator.
- To programme SaCas9, you need to design guide RNA (tracrRNA [2], 21 bp spacer flanked by direct repeats [2] ).
Advantages of SaCas9 compared with the conventional Streptococcus pyogenes Cas9:
- Smaller size (1053 amino acids against 1368) resulting in an easier expression/delivery
- Different PAM sequence recognised (5’-NNGRRT-3’ ) increasing the usability
- Higher efficiency of SaCas9 over SpCas9 [2]
[1] Ran, F. A., Cong, L., Yan, W. X., Scott, D. A., Gootenberg, J. S., Kriz, A. J., Zetsche, B., Shalem, O., Wu, X., Makarova, K. S., Koonin, E. V. Sharp, P.A., Zhang, F. 2015. In vivo genome editing using Staphylococcus aureus Cas9. Nature. 520 (7546). pp.186-191.
[2] Friedland AE, Baral R, Singhal P, et al. Characterization of Staphylococcus aureus Cas9: a smaller Cas9 for all-in-one adeno-associated virus delivery and paired nickase applications. Genome Biology. 2015;16:257. doi:10.1186/s13059-015-0817-8.
Please include a table of all the parts your team has made during your project on this page. Remember part characterization and measurement data must go on your team part pages on the Registry.
