...using bacterial phage vectors with CRISPR/cas9 sensitizing gene cassettes. Thse vectors are engineered to... tetracycline cas9 inducible knock-out system. His lab is also exploring the use of cas9 proteins from...were identified for 61 such diseases. Now single gene alleles have been identified for 4,163, over 50%...Lowe talked about his work on the 500+ human tRNA genes and, until now, we've overlooked regulatory fragments...may be involved in preventing silencing of self-genes. Dr. Mello presented a hypothesis that these pathways...involved in RNA based surveillance of repetitive gene families and provide a way to guide inheritance ... to replace the drug resistantance genes in the bacteria thus making the bacteria sensitive to known antibotics...

...can knock down genes by feeding worms bacteria expressing dsRNA or use CRISPR/Cas9 gene editing to easily...briggsae , etc.) gene or sequence: ID Plasmid Gene/Insert PI Don’t see your gene listed? Know of some great... via a CRISPR-Cas9 system. Cloning-free CRISPR for C. elegans , which uses purified Cas9, such as that...Tagging of Endogenous Genes with SapTrap Even more elegant: Single injection CRISPR/Cas9 in C. elegans External...Custom Genetic Modifications with CRISPR/Cas9 in Caenorhabditis elegans. Mos1 Based Transgenesis miniMos... used in worms, search Addgene's entire collection. ID Plasmid Description Gene/Insert PI Plasmids Expressing...which diminishes epigenetic silencing in the germline. A "FLP-Out" system for controlled gene expression in...

...engineered Cas9 variant might work for your experiment. PITChing MMEJ as an Alternative Route for Gene Editing...Corn’s lab describe how a better understanding of Cas9 biophysics can help enhance HDR. CRISPR 101: Ribonucleoprotein...tool to help you design, pick, and close guides for Cas9 and Cpf1. This tool supports over 250 genomes, and...template to increase efficiency of insertion and gene replacement. Care to share your favorite CRISPR ...Blugene and I represented Addgene at the recent Keystone meeting on Precision Genome Editing with Programmable...hopes to evaluate genome editing pipelines and generate a common lexicon for genome editing studies. These...desired edit, and I want to make sure that I (and Addgene) don’t forget this in the future. Here’s a list...

...the CRISPR-Cas9 system. Mehmet Fatih Bolukbasi from Scot Wolfe's lab described a chimeric Cas9 that is improving...CRISPR-Cas9 field – Jennifer Doudna, Feng Zhang, and Keith Joung. All Addgene depositors! Addgene was mentioned...versions of each, and Tim Lu even compared his "memory" gene codes to a Turing machine (also discussed in the...harnessed. Sharing CRISPR, synbio, & more via Addgene Addgene's addition to the poster session was a first...attendant called them, "long, skinny things") and generally exuding a very-tired-but-very-excited attitude...happens): Science is hard. Chris Voigt, another Addgene depositor, discussed some of the many challenges...That so many scientists are tackling this type of genetic engineering is a testament to how amazing the downstream...

... your gene of interest. For ideal use, this gRNA should be cloned into a vector containing Cas9 and the...et al. (2016).  “MMEJ-assisted gene knock-in using TALENs and CRISPR-Cas9 with the PITCh systems.” Nat ...harnessed MMEJ to create a new method for CRISPR gene knock-in, termed PITCh (Precise Integration into Target ...than that observed with NHEJ. PITCh: Using MMEJ for gene knock-in Building on the lab’s previous work, Sakuma...plasmic is cotransfected with a plasmid carrying Cas9, the PITCh-gRNA, and the locus-specifc gRNA. The...Contransfect the PITCh vector with the vector carrying Cas9 and the PITCh- and locus-specific gRNAs Step 4: ... is if the GFP-Puro will alter expression of the gene it follows. For increased versatility, it would ...

...-invasive systemic gene delivery in non-human primate (NHP) brains.  Systemic gene delivery capsids previously...Monomeric StayGold variants AAV.CAP-Mac for systemic gene transfer in primate brains Endothelial-specific ...peptide” that facilitates endosomal escape. Combining Cas9-PAGE with lentiviral sgRNA expression yielded editing...lines and primary cells. Although cell entry of a Cas9 ribonucleoprotein (RNP) was limited (likely due ...multiplexed guides. Figure 1: A) Cell-penetrating Cas9, fused to HIV TAT, Myc and SV40 Nuclear Localization...Localization Signals, and GFP. B) Workflow to assess Cas9-PAGE editing with viral sgRNA targeting mCherry reporter...Adeno-associated viral vectors for functional intravenous gene transfer throughout the non-human primate brain....

...CRISPR/Cas9 assembly for fast generation of multiple Arabidopsis mutants - Niko Geldner. CRISPR-Cas9 vector... an intronized Cas9 series of vectors and DNA modules for assembly of CRISPR/Cas9 constructs - Sylvestre... a new window) - Tools for gene and protein expression and mapping of gene expression in plants. Figshare...MoClo-compatible zCas9i Cloning Kit includes a high efficiency intron-optimized SpCas9-coding gene, zCas9i, and an...from genome engineering and gene regulation to fluorescent proteins. Addgene’s collection includes plasmids...plasmids containing promoters that drive general and species-specific gene expression (see the column ‘promoter...vectors: A modular plasmid assembly kit for multigene expression, gene silencing, and silencing rescue in plants...

...are created by fusing Cas9 nickase or catalytically inactive “dead” Cas9 (dCas9) to a cytidine deaminase...editors are available with multiple Cas9 variants including high fidelity Cas9’s. Further advancements have ... ID Plasmid Gene/Insert Promoter Selectable Marker PI Publication Bacteria ID Plasmid Gene/Insert Promoter...Plant ID Plasmid Gene/Insert Promoter Selectable Marker PI Publication Yeast ID Plasmid Gene/Insert Promoter...AID-mediated mutagenesis (TAM) from the Chang lab and CRISPR-X from the Bassik lab , in which Cas9 is fused...base editors, the evolved TadA domain is fused to a Cas9 protein to create the adenine base editor. Both ...Selectable Marker PI Publication Zebrafish ID Plasmid Gene/Insert Promoter Selectable Marker PI Publication...

... quantify Cas9 cleavage, thus exhibiting its versatile utility in quantifying multiple gene editing activities...reporter for rapid quantification of base-editing and Cas9 cleavage activity Article contributed by Angela ...lentiviral vector system that monitors both APOBEC- and Cas9-mediated Editing (ACE) in real-time. The reporter...Wanisch  Listen to the optogenetics podcast segment LOVTRAP is an optogenetic system that was recently...Harris lab showed that human APOBEC3A-Cas9n-UGI and APOBEC3B-Cas9n-UGI base editing complexes are more ... the University of North Carolina (plasmids at Addgene). It consists of two domains, LOV2 and Zdk1, where...function is more specific and much faster than any genetic or pharmacological manipulations on cells, and ...

... In Resistance Co-expressed Cas9 Depositing lab Cas9 species = S. pyogenes (PAM = NGG) 42875 pCRISPR BsaI...repair (HDR). ID Plasmid Gene/Insert Promoter PI Publication Prime Edit Cas9 H840A nickase fused to a ...dead dCas9, or dCas9 fused to a transcriptional repressor peptide like KRAB, can knock down gene expression... ID Plasmid Gene/Insert Promoter PI Publication 64325 3xFLAG-dCas9/p-bacteria 3xFLAG-dCas9 pLtetO-1 Fujii...template. ID Plasmid Gene/Insert Promoter PI Publication Activate Catalytically dead dCas9 fused to a transcriptional...transcription of a specific gene. Design your gRNA sequence to direct the dCas9-activator to promoter or ... plasmid to target the dCas9-activator to your specific locus. ID Plasmid Gene/Insert Promoter PI Publication...

...Delivery Resistance Co-expressed Cas9 Depositing lab Cas9 species = S. pyogenes (PAM = NGG) 51306 pUC57-Simple-gRNA...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...faithfully introduce specific genomic changes. ID Plasmid Gene/Insert Promoter PI Publication Empty gRNA Expression...backbone T7 BsaI In vitro transcription none, need Cas9 plasmid Chen Do you have suggestions for other plasmids...Transcriptional Regulation Activate Interfere Epigenetics RNA Targeting RNA Targeting RNA Editing Other...Other Applications Purify Tag Visualize dCas9-FokI Screen Pooled Libraries gRNAs Empty gRNA Vectors gRNAs...

...replace” gene editing method in which a reverse transcriptase (RT) is fused to the C terminus of Cas9 H840A... ID Plasmid Gene/Insert Promoter Selectable Marker PI Publication Bacteria ID Plasmid Gene/Insert Promoter...ID Plasmid Gene/Insert Promoter Selectable Marker PI Publication Drosophila ID Plasmid Gene/Insert Promoter...contains Cas9. ID Plasmid Expression System Promoter Guide RNA Type Cloning Enzyme Co-expressed Cas9 Selection...directs the Cas9 nickase to the target sequence where it nicks the non-target strand and generates a 3’ flap...nuclear localization signals, and two mutations in Cas9 to enhance nuclease activity; can be used in PE2...PE6 - small prime editors with optimized RT and/or Cas9 domains PE7 - addition of RNA-binding exonuclease...

...possible deletion strategies for genes and non-coding elements. For creating a gene knockout, two sgRNA located...evaluate the RNA, perform RT-qPCR for gene expression of the relevant gene 7,8 . To evaluate the protein, perform...Article Citation Generation of Genomic Deletions in Mammalian Cell Lines via CRISPR/Cas9. Bauer DE, Canver... CRISPR/Cas9 is an inexpensive, facile, and efficient genome editing tool that allows genetic perturbation...perturbation of genes and genetic elements. Here we present a simple methodology for CRISPR design, cloning,...for efficient loss-of-function studies of genes and genetic elements in mammalian cell lines. Protocol...excerpted from: Generation of Genomic Deletions in Mammalian Cell Lines via CRISPR/Cas9. Bauer DE, Canver...

...explains how gene drives work and what work he's planning on doing with CRISPR-based gene drives focusing...member of the Addgene Scientific Advisory Board with a particular interest in generating Cas9 based tools...Additional Resources on the Addgene Blog Learn More About Alex's Work with Cas9 Activators Michael Koeris...developing new CRISPR tools, and the potential uses of gene drives. Listen here! Podcast breakdown (Time : ...- 31:25 : Nature's defenses against gene drives. 31:26 - 33:08 : Plans for future work at the Wyss Institute... gene drives...became a member of the Addgene advisory board where he will help shape Addgene's future. Listen to this...

...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...for the EGFP Reporter Gene Guide RNA expression plasmids for endogenous human genes Guide RNA expression...these vectors for efficiently modifying endogenous genes in zebrafish (Hwang and Fu et al., Nat Biotechnol...expression plasmids for endogenous zebrafish genes... (AsCpf1 and LbCpf1) for use in mammalian cells SpCas9 variants with no detectable off-target effects ...

...multiple terminators can serve to interrupt genes (or gene clusters) in metabolic engineering. So far,...consists of one plasmid that expresses the S. pyogenes Cas9, a temperature-sensitive replicon for self-curing...pTargetF with the ccdB gene containing BsaI restriction enzyme sites at its ends to generate pEcgRNA. The ccdB...to successfully delete genes of interest in BL21(DE3) using the newly generated pEcCas/pEcgRNA system....used MUCICAT to optimize the gene dosage of glucose dehydrogenase and generated a strain that could achieve...J, Yang S (2015) Multigene Editing in the Escherichia coli Genome via the CRISPR-Cas9 System. Appl Environ...-curing, an arabinose inducible lambda-red gene, and an IPTG-inducible sgRNA sequence targeting the pMB1...

...CRISPR/Cas9 methodology with T cell receptor engineering (Ref 5)  as described by Blue Flame Addgene depositor...using randomly integrating viral vectors, CRISPR/Cas9 genome editing allows targeted engineering of the.... "Targeting a CAR to the TRAC locus with CRISPR/Cas9 enhances tumour rejection." Nature 543.7643 (2017...Additional Resources on the Addgene Blog Read Additional Cancer Blog Posts Addgene's Cancer Collection Pages...this multi-pronged attack. Inflammation and tumorigenesis The conference opened with a very interesting... between tumor and immune cells in real time. Addgene depositor Professor Max Krummel from UCSF showed...poster is pTomo-Ras which has been deposited at Addgene was used to produce the glioblastoma model for ...

...zebrafish gene of interest? Search Addgene's collection for plasmids that contain a zebrafish gene or sequence...Lab CRISPR/Cas9-based conditional mutagenesis in zebrafish - Wenbiao Chen Lab A CRISPR/Cas9 Vector System... zinc finger nucleases, TALENS, and CRISPR/Cas9-based gene editing make the zebrafish a stalwart research...zebrafish-specific gene expression database that provides information about when genes are active in developing...developing embryos. zfRegeneration - A database for gene expression profiling during regeneration. Seurat - R ...-type specific conditional gene inactivation - Jeffrey Essner Lab GeneWeld Vectors for Targeted Integration...modified CRISPR-Cas9 system - Shuo Lin Lab Re-purposed Ac/Ds transgenic system enabling CRISPR/dCas9-interference...

...CRISPR/Cas9 genetic circuits References: Nielsen AA, Voigt CA. Multi-input CRISPR/Cas genetic circuits...binding proteins with Cas9 guided by different promoters. The challenge in building genetic circuits is getting... to turn on genes at different times. It requires more complex synthetic networks. Addgene: What hurdles... that stop it from being used. One thing is that Cas9 is toxic. When cells carry CRISPR circuitry, it ...on an engineering project including hundreds of genes and regulation and resource control - everything...gives us enough hardware to control hundreds of genes. We’ve created enough regulatory elements, CRISPRs...the scale of genetic engineering, with the ultimate objective of genome design.” It’s genetic engineering...

...the Addgene blog CRISPR 101: Which Cas9 do I choose for my CRISPR experiment? CRISPR 101: Cas9 The ...long ago, spCas9 was the only Cas enzyme widely available and applied by researchers for gene targeting...they're available through Addgene. So whether you're looking for the exact Cas9 plasmid used by Jinek et...Resources on Addgene.org CRISPR software and resources Addgene CRISPR Guide Addgene’s CRISPR 101 eBook...and trans activity of each entry. For example, SpyCas9a (one of the most frequently used Cas enzymes) ... trans activity. AsCas12a, on the other hand, generates staggered DNA nicks at its target in cis and cleaves...end goal. CasPEDIA also features useful links to Addgene resources throughout the site. In the Experimental...