...that do not co-express a Cas protein can be paired with a wide variety of Cas-containing plasmids. Do ... expression systems. Cut Fully functional CRISPR/Cas enzymes will introduce a double-strand break (DSB...Promoter Selectable Marker PI Publication Nick CRISPR/Cas nickase mutants introduce gRNA-targeted single-strand...of the double-strand breaks created by wild type Cas enzymes. To use a nickase mutant, you will need two...When using CRISPR, you will need to express both a Cas protein and a target-specific gRNA in the same cell...time. Single plasmids containing both the gRNA and Cas protein act as all-in-one vectors, but their function...PI Publication Interfere Catalytically dead dCas9, or dCas9 fused to a transcriptional repressor peptide...

...that do not co-express a Cas protein can be paired with a wide variety of Cas-containing plasmids. ID ...When using CRISPR, you will need to express both a Cas protein and a target-specific gRNA in the same cell...time. Single plasmids containing both the gRNA and Cas protein act as all-in-one vectors, but their function...Enzyme(s) Delivery Resistance Co-expressed Cas9 Depositing lab Cas9 species = S. pyogenes (PAM = NGG) 51306...Editing Other Applications Purify Tag Visualize dCas9-FokI Screen Pooled Libraries gRNAs Empty gRNA Vectors...designed for use in Xenopus. Cut Fully functional Cas9 enzymes designed to introduce a double-strand break...surrounding the DSB is introduced along with the Cas9 and gRNA plasmids, the cell may instead repair the...

...that do not co-express a Cas protein can be paired with a wide variety of Cas-containing plasmids. ID ...designed for use in plants. Cut Fully functional CRISPR/Cas enzymes will introduce a double-strand break (DSB...Promoter Selectable Marker PI Publication Nick CRISPR/Cas nickase mutants introduce gRNA-targeted single-strand...of the double-strand breaks created by wild type Cas enzymes. To use a nickase mutant, you will need two...When using CRISPR, you will need to express both a Cas protein and a target-specific gRNA in the same cell...time. Single plasmids containing both the gRNA and Cas protein act as all-in-one vectors, but their function...PI Publication Interfere Catalytically dead dCas9, or dCas9 fused to a transcriptional repressor peptide...

...RNA-guided endonuclease) Tools: Cas-OFFinder & Cas-Designer From the Kim Lab , Cas-OFFinder identifies gRNA ...of off-target effects. Cas-Designer integrates information from the Kim Lab's Cas-OFFinder and Microhomology... sativa), tomato, corn, monkey (macaca mulatta). Cas-Designer searches for targets that maximize knockout...tropicalis ), and zebrafish. Off-Spotter: tool for CRISPR/Cas design Program for designing optimal gRNAs. Provides... potential DNA target sites for TALEs and CRISPR/Cas. CRISPRdirect From the Database Center for Life Science...vectors using ssDNA oligos p201G Cas9 ; p201B Cas9 ; p201H Cas9 ; p201N Cas9 ; pUC gRNA Shuttle PDF 488.6 ...CRISPR RNA array: Cas9 (pX260) or Cas9 D10A (pX334) ; tracrRNA: Cas9 (pX330) or Cas9 D10A (pX335) ; PDF...

...Species of Cas enzyme and gRNA Species of promoter and expression pattern of promoter for Cas enzyme and...compatible with smaller Cas enzymes) Transient or stable expression of SaCas9 and/or gRNA Infects dividing...sequence, you may want to consider alternative Cas enzymes ( see Cas9 variants and PAM sequences ). Once possible... exon or essential protein domains High-fidelity Cas enzymes increase specificity. Dual-nickase approach...of System Application Mammalian expression vector Cas enzyme promoter can be constitutive (CMV, EF1alpha...specificity, you can also consider using a high-fidelity Cas enzyme. In addition to off-target activity , it is...Once you have successfully delivered the gRNA and Cas enzyme to your target cells, it is time to validate...

...more information on the wide variety of Cas enzymes? CasPEDIA is an encyclopedia of Class 2 CRISPR systems...Protocols gRNA Design Tools CRISPR Blog Posts CRISPR/Cas nickase mutants introduce gRNA-targeted single-strand...of the double-strand breaks created by wild type Cas enzymes. To use a nickase mutant, you will need two...Editing Other Applications Purify Tag Visualize dCas9-FokI Screen Pooled Libraries gRNAs Empty gRNA Vectors...

...proteins. This CRISPR/Cas plasmid-system consists of two components: A vector containing Cas9 and a validated...marker. The first deposited plasmids in this CRISPR-Cas tagging system were tested by tagging transcription...into human FBL locus Jorgensen Lab SapTrap CRISPR/Cas Toolkit SapTrap is a modular toolkit from Erik Jorgensen's...GFP targeting vector. Jorgensen Lab SapTrap CRISPR/Cas Toolkit Kanemaki Lab Auxin-Inducible Degron Tagging...Kanemaki's lab has developed a simple and scalable CRISPR/Cas-based method to tag endogenous proteins by using ...Auxin-inducible degron (AID) tagging with CRISPR/Cas to generate conditional alleles of essential nuclear...Doyon lab, eSpCas9(1.1)_No_FLAG_AAVS1_T2 (Addgene #79888) , which expresses an untagged Cas9 AND a gRNA...

... CRISPR dCas9-FokI CRISPR: dCas9-FokI...Tools CRISPR Blog Posts A catalytically inactive Cas9 (dCas9) is fused to FokI nuclease. When FokI dimerizes...with dCas9 fused to FokI for creating double-strand breaks (DSBs). CRISPR...Editing Other Applications Purify Tag Visualize dCas9-FokI Screen Pooled Libraries gRNAs Empty gRNA Vectors...gRNAs, binding ~15-25 bp apart, are required for dCas9-FokI to dimerize at a given region of the genome...

...Engineering Cut Wild-type Cas enzymes generate double strand breaks (DSBs). Nick Mutated Cas nickase enzymes generate... generate single-strand DNA breaks. Prime Edit Cas nickase enzymes fused to reverse transcriptase install... without promoting DSB formation. Delete NEW Cascade-Cas3 generates large genomic deletions. Insert NEW...Regulation Activate dCas9 fused to an activator peptide activates gene expression. Interfere dCas9 alone, or fused...repressor peptide, inhibits transcription. Epigenetics dCas9 fused to an epigenetic modifier alters chromatin..., not DNA, across multiple systems. RNA Editing Cas13 enzymes fused to deaminases can edit bases in RNA...powerful tool for genome-wide screening. Purify dCas9 fused to an epitope tag can be used to isolate a...

...for using iSPI, rEcoli XPS, and pSerOTS-C1* (V70) (ca. 2022) from the Rinehart lab: Methods for iSPI, rEcoli...PubMed (Link opens in a new window) Improved Reagents (ca. 2015) described in: A flexible codon in genomically...window) Protocol for using the Improved Reagents (ca. 2015) from the Rinehart lab: Users Guide for Improved...Phosphoprotein Reagents (DOCX, 525.9 KB) Reagents (ca. 2014) described in: Enhanced phosphoserine insertion...Link opens in a new window) Protocol for Reagents (ca. 2014) from the Rinehart lab: Users Guide for 2014...Phosphoprotein Reagents (PDF, 691.3 KB) Original Kit (ca. 2011) described in: Expanding the genetic code of...