RGENs (RNA Guided Endonucleases)
RNA-Guided ENdonucleases (RGENs) are novel, programmable genome engineering tools that are adapted from CRISPR/Cas system. Cas9 forms a sequence-specific endonuclease when complexed with two RNAs, one of which guides target selection (crRNA) and and the other is a constant component (tracrRNA).
RGENs recognize a target specific sequence that is 23bp in length, ending with two guanines (GG). This minimal requirement for target site selection means that RGENs can be designed to cleave any region of the genome (every 8bp in theory).
RGENs have recently been shown to support efficient genome engineering in human cells, zebrafish and bacteria. In addition, the use of RGENs allows multiplexing of genome engineering processes (e.g., knockout of multiple genes in a cell at one time).
RGEN also shows impressive sequence specific activity. Activity and specificity of RGEN is comparable to TALEN.
HEK293 cells are transfected with increasing amount of plasmid expressing sgRNA in the presence or absence of Cas9 plasmid. In/del caused by CRISPR/Cas9 complex was assessed by T7E1 assay.
|We provide custom CRISPR/Cas9 (RGEN) synthesis service with or without validation.|
IVT sgRNA Synthesis Service (aRGEN)
sgRNA Vector Synthesis Service (dRGEN)
sgRNA Validation Service
Guide RNA (gRNA) Design
|In general, find gRNA sequence with no off-targets for 1 bp and 2 bp mismatches, and contains 45~70% GC. If you cannot avoid off-targets, mismatches located near PAM are preferred over the ones at 5' side of gRNA. Please note that non-canonical PAM (-NAG, -NGA) can also be cleaved. |
We have developed our gRNA Tool that outputs off-target sequences, GC content and other features. You can also use publically available tools to search with various options.
Cas-Offinder (to see the off-target list)
eGFP Control sgRNA
- Laminar flow downregulates Notch activity to promote lymphatic sprouting. Choi D et al. (2017) J. Clin Investigation. In press.
- Generation and characterization of tamoxifen-inducible Pax9-CreER Knock-In Mice using CrispR/Cas9. Jifan F et al. (2016) Genesis 54(9):490-496.
- Efficient delivery of nuclease proteins for genome editing in human stem cells and primary cells. Liu J et al. (2015) Nat Protoc. 10:1842-1859.
- Highly efficient RNA-guided genome editing in human cells via delivery of purified Cas9 ribonucleoproteins. Kim S et al. (2013) Gen Res. 24:1012-1019.
- Rapid and highly efficient mammalian cell engineering via Cas9 protein transfection. Liang X et al (2015) J Biotech. 208: 44-53.
|DR01||Custom sgRNA/Cas9 expression plasmids||Inquire|
|DR02||Activity validated sgRNA/Cas9 vector by mismatch sensitive nuclease assay in cells||Inquire|
|AR01||Custom IVT sgRNA, injection ready (50 ug)||Inquire|
|AR02||1~3 in vitro validated sgRNA, injection ready (50ug each)||Inquire|
|AR04||eGFP control sgRNA (20 ug or 100 ug)||Inquire|
|Surrogate reporter system|
|RV01||GFP surrogate reporter system / 5 ug||$350|
|RV02||MACS/GFP surrogate reporter system / 5 ug||$350|
|RV03||HygR/GFP surrogate reporter system / 5 ug||$350|
|CV01||Cas9 co-transfection vector (CMV promoter) / 5 ug||$350|
|CV02||Cas9/HygR/GFP vector (CMV promoter) / 5 ug||$350|
|CV03||Cas9/RFP/PuroR vector (CMV promoter) / 5 ug||$350|
|CV11||Cas9 co-transfection vector (EF1a promoter) / 5 ug||$350|
|CV12||Cas9/HygR/GFP vector (EF1a promoter) / 5 ug||$350|
|CV13||Cas9/RFP/PuroR vector (EF1a promoter) / 5 ug||$350|