...resources include plasmids expressing Cas9, empty gRNA backbones, and constructs for endogenous tagging. ... genes by feeding worms bacteria expressing dsRNA or use CRISPR/Cas9 gene editing to easily engineer specific...characterized which allow researchers to express desired constructs in precise cell types and/or developmental... via a CRISPR-Cas9 system. Cloning-free CRISPR for C. elegans , which uses purified Cas9, such as that...that expressed from pHO4d-Cas9 from Michael Nonet. Protocol can be found at the (Link opens in a new window...and spatial control of gene expression in C. elegans by combining expression of the FLP recombinase from...Worm Plasmids and Resources Plasmid Tools Worm Constructs Kits and Collections Other Resources The nematode...

...full selection of plasmids expressing different functional variants of Cas9 and empty gRNA backbones. ... assembly of one or more gRNA expression cassettes - Qi-Jun Chen. Cas9 and gRNA backbone for use in rice...DNA modules for assembly of CRISPR/Cas9 constructs - Sylvestre Marillonnet and Johannes Stuttmann. Combined...CRISPR/Cas9 assembly for fast generation of multiple Arabidopsis mutants - Niko Geldner. CRISPR-Cas9 vector...creating plant expression vectors: A modular plasmid assembly kit for multigene expression, gene silencing...create plant expression vectors containing several cassettes and generate multi-construct transgenic plants... - Tools for gene and protein expression and mapping of gene expression in plants. Figshare - Plant Illustrations...

...μg of each CRISPR/Cas9 construct (10 μg total). Add 0.5 μg of GFP expression construct. Electroporate cells...markers, respectively, or constructs in which multiple sgRNAs may be expressed from a single plasmid. Add...DNA yield. Perform maxi-prep for each CRISPR/Cas9 construct. Transfecting CRISPRs into Cells of Interest...cells that received high levels of the CRISPR/Cas9 constructs. Plate sorted cells individually into 96-well... for cloning and expression purposes. NOTE: Here we discuss the usage of the pSpCas9(BB) plasmid (pX330...plasmid allows for the simultaneous expression of sgRNA and SpCas9, but does not contain markers for selection...genomic deletions in mammalian cell lines using CRISPR/Cas9 CRISPR...

...inactive bacterial Cas9 expression plasmid 44250 pwtCas9-bacteria A wild-type Cas9 expression plasmid for use... gRNA expression plasmid for use with either human codon-optimized Cas9 constructs 44249 pdCas9-bacteria...customizable gRNA expression plasmid for use with the bacterial Cas9 constructs 46910 pHR-SFFV-dCas9-BFP Human...Description 44246 pdCas9-humanized A catalytically inactive, human codon-optimized Cas9 expression plasmid 44247...pHR-SFFV-dCas9-BFP-KRAB Human expression vector containing SFFV promoter, dCas9 that is fused to 2x NLS, tagBFP...46912 pMSCV-LTR-dCas9-VP64-BFP Human expression vector containing MSCV LTR promoter, dCas9 that is fused...and tagBFP 51023 pSLQ1658-dCas9-EGFP Human expression vector containing dCas9 that is fused to 2x NLS and...

...conversion to a dual guide construct. Plasmids 175573 and 175574 express dCas9-KRAB-MeCP2 for repression... and 159655 are single guide expressing double stranded break constructs. Plasmids 171098 and 175570 allow...U6-guide cassettes to make dual guide expressing plasmids dCas9-KRAB-MeCP2 fusion for gene and non-coding...enhancer sequence repression dCas9-VPR fusion for gene activation CRISPR/Cas9 Plasmid Variants Figure 1:...repression. Plasmids 175571 and 175572 express dCas9-VPR for activation. Usage Guide sequences can be designed...in retinal development the lab utilizes a CRISPR/Cas9 based approach to knockout, repress, and activate...taken advantage of the Zhang lab all-in-one (guide/Cas9/reporter) pX458 plasmid family. They have made several...

...Plasmids: pX330S-2-PITCh - expresses Cas9 & PITCh-gRNA pX330A-FBL/PITCh - expresses Cas9, PITCh-gRNA, & FBL-specific...Doyon lab, eSpCas9(1.1)_No_FLAG_AAVS1_T2 (Addgene #79888) , which expresses an untagged Cas9 AND a gRNA... that uses PCR to generate both an expression cassette for the Cas9-programming sgRNA and HR donors for...: 3xFLAG-2xSTREP Construct for integrating at the AAVS1 "safe harbor" locus: eSpCas9(1.1)_No_FLAG_AAVS1...has been adapted to allow tagging of proteins expressed from their natural chromosomal context. These ... consists of two components: A vector containing Cas9 and a validated gRNA, based on the Zhang lab's PX458...need to design and clone in a gRNA(s) to guide the Cas9 protein to your target sequence, as well as design...

...specifity of Staphylococcus aureus Cas9 (SaCas9 KKH variant) Engineered CRISPR-Cas9 nucleases with altered PAM...the Cas9 endonuclease) that cleaves the target DNA. The Joung lab recently described gRNA and Cas9 expression...design of short oligonucleotides needed to construct gRNA expression vectors. More information about RGNs and...) CRISPR-Cas/RGN expression plasmids for use in human cells CRISPR-Cas/RGN expression plasmids for use...zebrafish Guide RNA expression plasmids for the EGFP Reporter Gene Guide RNA expression plasmids for endogenous...reagents: Cpf1 expression plasmids (AsCpf1 and LbCpf1) for use in mammalian cells SpCas9 variants with ...endogenous human genes Guide RNA expression plasmids for endogenous zebrafish genes...

... EGFP, and sgRNA expression hSyn AAV Zhang 60820 pSECC Cre, Cas9 and sgRNA expression EF-1 alpha Lentiviral...Livet loxP (floxed) Constructs These constructs allow for Cre-regulated gene expression. Depending on the...the construct, Cre may activate or repress gene expression. Common construct types include: Cre-dependent...shRNA Expression: Cre-lox can be used to turn shRNA constructs on or off. In floxed-shRNA constructs, Cre...return gene expression to physiological levels. In lox-STOP-lox shRNA constructs, Cre expression promotes...Cre-dependent Cas9: Cre-dependent Cas9 can be knocked in to a mouse at the Rosa26 locus to facilitate Cas9 mediated...have developed constructs to activate/inactivate genes when Cre is present. By expressing Cre at specific...

...dual-dCas9VP64-sgExpression : Dual expression construct expressing both dCas9VP64 and sgRNA from separate... pAC153-dual-dCas9VP96-sgExpression : Dual expression construct expressing both dCas9VP64 and sgRNA from...pAC154-dual-dCas9VP160-sgExpression : Dual expression construct expressing both dCas9VP64 and sgRNA from...Dual expression construct expressing both dCas9VP48 and sgRNA from separate promoters. 48238 pAC152-dual-dCas9VP64...pmax expression vector. 48224 pAC92-pmax-dCas9VP96 : dCas9VP96 on pmax expression vector. 48225 pAC93-pmax-dCas9VP160...produced by PCR), or expressed from a single dual-expression vector [48236~48240]. dCas9 activator ORFs (with...48223~48227] + sgRNA expression vector or the linear DNA containing the sgRNA expressing fragment produced...

...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...and selection for Cas9-triggered homologous recombination Cas9-sgRNA target construct ; pMA122 (negative...FLAG tagged dCas9 PDF 107.4 KB Goldstein Nematode: gRNA design and cloning Cas9-sgRNA construct PDF 355.3...putative target sites for S. pyogenes Cas9, S. thermophilus Cas9, or Cpf from your input sequence or list....2 KB Church gRNA design and cloning for Cas9 orthologs Cas9 plasmids PDF 104.1 KB Chen and Wente Zebrafish...vitro transcription, injection gRNA core ; Cas9 ; optimized Cas9 PDF 66.8 KB Fujii gRNA design and cloning...

...double nickase, you could express two spacers and use PX335 to express the Cas9n (nickase). The concept ...need to have four spacers co-expressed and using PX335 to express the Cas9n (nickase). Homologous Recombination...nickase system is that you can express two different chimeric gRNAs with the Cas9 nickase which will together...the Cas9 nickase doesn't have the ability to induce double-stranded breaks like the wildtype Cas9 does...start site to have high expression, so adding this G could help with expression, though it is possible ...shows the construct from our 'Split RNA' design, where the pre-processing form of crRNA is expressed and a...of the target spacer, but then when expressed in cell and with Cas9 and tracrRNA, it will be processed ...

... 52962 lentiCas9-Blast Sp Cas9 Cut Blasticidin Expresses human codon-optimized S. pyogenes Cas9 protein...backbone. Zhang Cas9 and Accessories for Activating Gene Expression Catalytically-dead Cas9 (dCas9) can be fused...Root Cas9 Lentivirus Cas9 viruses can be used to interrogate the genome in a variety of ways. Cas9 proteins...plasmids resource page . Cas9 for Editing Genomic Sequences To perform genome edits, Cas9 nuclease is used to...Resistance Activity PI 61422 dCAS9-VP64_GFP dCAS9 (D10A, H840A) none Expresses dCAS9-VP64 activator with 2A ...Addgene's viral vector packaging service. Browse constructs currently available in lentiviral format. Viral...Browse In-Stock Lentivirus Pooled CRISPR Libraries Cas9 Pooled Barcoding Libraries Control Addgene's lentiviruses...

...resources for a full selection of plasmids expressing Cas9 and empty gRNA backbones. A few examples of... Using CRISPR/Cas9 - Jeffrey Essner Lab CRISPR-Cas/RNA-Guided Nuclease (RGN) expression plasmids - Keith...Lab CRISPR/Cas9-based conditional mutagenesis in zebrafish - Wenbiao Chen Lab A CRISPR/Cas9 Vector System...fusion expression in organisms or cells in which an exogenous mRNA can be injected and expressed. EXPRESs...in zebrafish. Zebrafish expression atlas - A zebrafish-specific gene expression database that provides ...modified CRISPR-Cas9 system - Shuo Lin Lab Re-purposed Ac/Ds transgenic system enabling CRISPR/dCas9-interference...morpholinos, zinc finger nucleases, TALENS, and CRISPR/Cas9-based gene editing make the zebrafish a stalwart...

...the Doudna Lab . Plasmid Expression System Cloning Enzyme Co-expressed Cas9 Cas9 System Selection PI pCRISPR...Mammalian Lentiviral expression of Cas9, dCas9, or dCas9-VP64 along with 1-4 sgRNAs expressed from independent... plasmid contains Cas9 and if so, which function of Cas9. We have plasmids with Cas9 that can cut, activate...be used to express multiple gRNAs from a single plasmid with or without expression of Cas9. Highlighted...system for constructing all-in-one expression vectors containing multiple guide RNA expression cassettes.... Can express Cas9 with up to 3 gRNAs. Frew pSQT1313 Mammalian Single plasmid for the expression of multipe...cassettes are assembled into a Cas9 expressing vector. Yang A Robust CRISPR/Cas9 System for Convenient, High-Efficiency...

...Technique PI Purpose pCas9 42876 CRISPR Luciano Marraffini Bacterial expression of Cas9 nuclease gRNA. For...so multiple constructs can be made and tested at once. Return to Top Controlled Expression It isn’t always... constitutively express your gene of interest at high levels. Perhaps high expression leads to slow bacterial... on expression only at a specific time. This collection has a variety of plasmids whose expression levels...Anhydrotetracycline inducible expression of catalytically inactive Cas9 ( S. pyogenes ) which can be combined... bacterial expression systems to find out more! Genome Engineering Although overexpressing or knocking...see here . pwtCas9-bacteria 44250 CRISPR Stanley Qi Anhydrotetracycline inducible expression of wild-type...

... while also adding additional filters such as expression, vector type, species, and more. Browse Browse...for Golden Gate assembly. MoClo Toolkit Plant Expression Sylvestre Marillonnet 95 plasmids for cloning...standardized parts to create eukaryotic multigene constructs. Pooled Libraries Pooled Library name Type PI...Stiewe Use these donor vector libraries for CRISPR/Cas9-mediated saturation genome editing of human TP53...

...plasmids expressing HTT, C9ORF72, LRRK2, Synuclein, and more. New and Noteworthy: Use tau constructs to study...Neurodegeneration Plasmid Collection A collection of plasmids expressing wild type or mutant ORFs, gene fragments, and...Target neural oxytocin receptors using an AAV-CRISPR/Cas9 strategy for gene editing across divergent rodent...species. Boender et al. Sci Adv. 2023 Jun 2. Use Cas9 in astrocytes. Endo et al Science. 2022 Nov 4. Base... dopamine research. CBA-driven, Cre-dependent expression of diphtheria toxin receptor (DTR)–GFP fusion...2023 Dec 26. Induce CNS cells with concomitant expression of aggregation-prone proteins with the iPSC toolbox...Dec 2. Use PiggyBac plasmids with tet-inducible expression of transcription factors for sensory, motor, ...

... available with multiple Cas9 variants, including high fidelity Cas9s, Cas9s with various PAM requirements...to either co-infect with a Cas9-producing lentivirus or use a Cas9-expressing stable cell line. After applying...strand SpCas9-HF1 - disrupt Cas9’s interactions with DNA phosphate backbone HypaCas9 - increase Cas9 proofreading...new tools. Figure 3: Overview of Cas9n Nickase and dead Cas9 The two Cas9 nuclease domains, RuvC and HNH...enhanced specificity by engineering Cas9 into high fidelity Cas9s (hfCas9) . There is currently no defined...application. Cas9 Orthologs While SpCas9 is the most commonly used Cas enzyme, additional Cas9 orthologs ...Additionally, constructs with Cas9 fused to proteins that modulate HDR or NHEJ components, like miCas9 , Cas9...