Source:
Generated By: https://synbiohub.org/public/igem/igem2sbol/1
Created by: Zexu Li
Date created: 2016-10-07 11:00:00
Date modified: 2016-10-09 12:04:45
Eukaryotic tCAS9
| Types | DnaRegion |
| Roles | Coding CDS |
| Sequences | BBa_K1982007_sequence (Version 1) |
Description
tCas9 is a standardized (RFC 10) protein.Interacting with a DNA-binding RNA and fused with different effector domains it can be used for specific gene regulation. Cas9 is the main protein of the CRISPR/Cas system II of Streptococcus pyogenes. CRISPR systems protect bacteria and archaea from phages by recognizing and cleaving of invading phage DNA.This recognition is based on Watson Crick base pairing between a short RNA, called crRNA, and the complementary DNA strand. A second RNA, called tracrRNA, connects crRNA and Cas9. These three parts together form a protein-RNA-DNA complex with the targeted DNA strand [1] Cas9 became of great interest for research concerning DNA targeting, because of its ability to recognize site specific DNA strands by a crRNA. At first the functionality of Cas9 was modified by exchanging aminoacids. As a result, Cas9 was able to introduce mutations within the genome of several organisms by causing double strand breaks [2][3]. Then, it was converted from a nuclease to a nickase introducing single strand breaks [4] and lately it was converted to an enzymatically inactive form, called dCas9 [5]. This tCas9 is standardized (RFC 10). It can be used as a DNA binding protein, that can be fused with different effectors in order to regulate gene expression.Notes
mutant Pst 1 :gctacctgcaG to gctacctgcaCmutant Pst 1 :actatctgcaG to actatctgcaC
Source
plasmid from GE Share company.[1] Westra E.R., Swarts D.C., Staals R.H., Jore M.M., Brouns S.J., van der Oost J. (2012). The CRISPRs, they are a-changin': how prokaryotes generate adaptive immunity. Annu Rev Genet. 46, 311-39
[2] Mali P., Yang L., Esvelt K.M., Aach J., Guell M., DiCarlo J.E., Norville J.E., Church G.M. (2013). RNA-guided human genome engineering via Cas9. Science 339(6121), 823-6
[3] Jiang W., Bikard D., Cox D., Zhang F., Marraffini L.A. (2013). RNA-guided editing of bacterial genomes using CRISPR-Cas systems. Nat Biotechnol. 31(3), 233-9
[4] Cong, L., Ran, F.A., Cox, D., Lin, S., Barretto, R., Habib, N., Hsu, P.D., Wu, X., Jiang, W., Marraffini, L.A., Zhang, F. (2013). Multiplex Genome Engineering Using CRISPR/Cas Systems. Science 339 (6121), 819-23
[5] Qi L.S., Larson M.H., Gilbert L.A., Doudna J.A., Weissman J.S., Arkin A.P., Lim W.A. (2013). Repurposing CRISPR as an RNA-guided platform for sequence-specific control of gene expression. Cell 152(5), 1173-83
| Sequence Annotation | Location | Component / Role(s) |
| Eukaryotic rbs atg | 1,18 19,21 | feature/rbs ribosome_entry_site feature/start start_codon |