gRNA#2 for SaCas9 targeting of IKKβ under U6 promoter
gRNA for SaCas9 targeting the second exon (genomic location: chr8:22706137) of mouse IKKβ (Inhibitor of nuclear factor kappa-B kinase subunit beta ) gene under the control of human U6 (RNA polymerase III) promoter.
This gRNA is a candidate for IKKβ gene knockout which will inhibit cytokine secretion in the NF-κB (nuclear factor kappa-light-chain-enhancer of activated B cells) pathway.
Usage and Biology
Guide RNA (gRNA) is a hundred base-long molecule, it includes a scaffold sequence which has a unique two dimensional structure necessary for Cas9 binding and a spacer sequence designed to target a specific location in the genome. gRNA scaffold sequence for SaCas9 was used.
In the NF-κB pathway, IKKβ phosphorylates the IkB inhibitory protein, a process which enables the release of NF-κB to the nucleus where it binds to its cognate DNA sequences to induce gene expression of several cytokines such as IL-1α and TNF. These cytokines have a crucial role in the induction of A1 reactive astrocytes. By knocking out the IKKβ gene we could reduce the IL-1α and TNF secretion from microglia cells and reduce the amount of reactive astrocytes by preventing the formation of their reactive form. By reducing the amount of reactive astrocytes we will prevent motor neuron death and prolong ALS patient survival.
1. Frakes, A. E., Ferraiuolo, L., Haidet-Phillips, A. M., Schmelzer, L., Braun, L., Miranda, C. J., ... & Popovich, P. G. (2014). Microglia induce motor neuron death via the classical NF-κB pathway in amyotrophic lateral sclerosis. Neuron, 81(5), 1009-1023.
2. Ran, F. A., Cong, L., Yan, W. X., Scott, D. A., Gootenberg, J. S., Kriz, A. J., ... & Koonin, E. V. (2015). In vivo genome editing using Staphylococcus aureus Cas9. Nature, 520(7546), 186.
3. Liddelow, S. A., Guttenplan, K. A., Clarke, L. E., Bennett, F. C., Bohlen, C. J., Schirmer, L., ... & Wilton, D. K. (2017). Neurotoxic reactive astrocytes are induced by activated microglia. Nature, 541(7638), 481.
Sequence and Features
- 10COMPATIBLE WITH RFC
- 12COMPATIBLE WITH RFC
- 21COMPATIBLE WITH RFC
- 23COMPATIBLE WITH RFC
- 25COMPATIBLE WITH RFC
- 1000COMPATIBLE WITH RFC