...fused to the N-terminus of LSSmOrange pLSSmOrange-C1 Orange Mammalian Express a gene of interest fused ... GFP variant) commonly used with mRuby2 pLSSmOrange-N1 Orange Mammalian Express a gene of interest fused...to measure protein interactions or conformation changes, as well as FRET standards. Plasmid...intermolecular or bimolecular FRET) or (2) conformational changes within a protein where the donor and acceptor are...fused to the C-terminus of LSSmOrange mRuby2 Red Mammalian Expresses mRuby2 (a RFP variant) commonly used ...

...of long-range inhibitory neurons. Preprint (Link opens in a new window) Yu K, Ahrens S, Zhang X, Schiff...ZR, Chen WZ, Liu MZ, Chen XJ, Wan L, Zhang XY, Yuan L, Lin JK, Wang M, Zhou L, Xu XH, Sun YG. 2019. Tac1...Haynes TM, Hochbaum DR, Huang KW, Hyun M, Umadevi Venkataraju K, Straub C, Wang W, Robertson K, Osten P... Perry C, Diester I, Boyce FM, Bass CE, Neve R, Huang ZJ, Deisseroth K. 2014. Targeting cells with single...AC, Tepler S, Poulin JF, Ansorge M, Awatramani R, Kang UJ, Rayport S. 2018. Dopamine neuron glutamate cotransmission...Neurons in Barrel Cortex Reveals Local and Long-Range Circuit Motifs. Cell Rep. 28(13):3450-3461. PubMed...

...University of California, Berkeley. CRISPR Software Sanger Indel Analysis ICE (Inference of CRISPR Edits) ... at a given locus. To use the tool, you'll need Sanger sequencing reads from PCR amplicons that cover ... budding and fission yeast. Target Finder (Feng Zhang lab) Identifies gRNA target sequences from an input...non-model invertebrate species. Forums and FAQs Feng Zhang lab: Tips for CRISPR design, homology directed repair...cloning . CRISPR Discussion Group (moderated by Feng Zhang lab): Forum/FAQ Drosophila: General Advice and Forum...pSL1180-HR-PUbECFP & pSL1180polyUBdsRED PDF 583.3 KB Zhang gRNA cloning CRISPR RNA array: Cas9 (pX260) or Cas9...Cas9 (pX330) or Cas9 D10A (pX335) ; PDF 248.6 KB Zhang sgRNA cloning lentiCRISPR v2 packaging plasmids:...

...material will be rearranged. The schematic below shows the three types of rearrangements: inversion, deletion...Cre-Lox system and how it can be used for genetic rearrangements and recombination. Use Addgene's curated tables...2A-Cre-WPRE-hGHpA-ITR Cre, luciferase, and sgRNA expression EFS AAV Zhang 60229 AAV:ITR-U6-sgRNA(backbone)-pCBh-Cre-WPRE-hGHpA-ITR...-hGHpA-ITR Cre and sgRNA coexpression Cbh AAV Zhang 60231 AAV:ITR-U6-sgRNA(backbone)-hSyn-Cre-2A-EGFP-...KASH-tagged EGFP, and sgRNA expression hSyn AAV Zhang 60820 pSECC Cre, Cas9 and sgRNA expression EF-1 ...Jacks 68468 Cas9-2A-Cre Cas9 and Cre CAG Mammalian Zhang 68477 pTRE:iRFP670-EFS:Cre-2A-GFP iRFP670, Cre, ...86641 pLenti-hSynapsin-Cre-WPRE Cre hSyn Lentiviral Wang 86794 pCL20c-MSCV-IRES-CRE Cre MSCV Mammalian Roussel...

... producing antibodies, can change the constant region, and therefore change the isotype of antibodies ...variable regions, shown in light blue and light orange, that bind to a specific antigen, triggering an...constant region, and is shown in dark blue and dark orange. The constant region is a stable part of the antibody... There are several alternatives to antibodies, ranging from antibody fragments (shown in Figure 2) to ...expensive than HRP, fluorophores are available in a range of colors activated by different wavelengths, allowing... assay run. The standard curve should contain a range inclusive of all expected protein concentrates in...to distinguish cell types. Multiplex assays can range from two antibodies to upwards of twenty. Laser ...

...via use of CRISPR-Cas9. Wu Y, Liang D, Wang Y, Bai M, Tang W, Bao S, Yan Z, Li D, Li J. Cell Stem...cells are used for DNA extraction, although a wide range of cell numbers is acceptable. The bulk sorted cells..., mini-prep each one, and subject each clone to Sanger sequencing to characterize each deletion allele...Ran FA, Hsu PD, Wright J, Agarwala V, Scott DA, Zhang F. Nature Protocols . 2013. PubMed . Characterization...colonies using the Qiagen Miniprep Kit. Zhang S, Cahalan MD. J. Vis. Exp . 2007. PubMed . Transformation...

...that selection is not required. pET-HIS-Sangamo 40786 TALEN Gang Bao Gateway cloning and expression of ...optimized FokI endonuclease domain. pET-Sangamo-His 40787 TALEN Gang Bao Gateway cloning and expression of...Various Plasmids Blue/UV , Cyan , Green , Yellow , Orange , Red , Far-Red , Near-Infrared , Large Stokes ...dissect an individual protein’s function under a wide range of highly controlled conditions in vitro , or, if..., Luciferase (LuxCDABE) Attila Karsi Broad host range vectors for expressing luciferase and fluorescent...

...Dillon CP, Kwiatkowski AV, Sievers C, Yang L, Kopinja J, Rooney DL, Zhang M, Ihrig MM, McManus MT, Gertler ...expression; Designed for inducing RNAi in a wide range of cell types, tissues and organisms Luk Parijs ...assessment of gene knockdown. Moore CB, Guthrie EH, Huang MT, Taxman DJ. Methods Mol Biol. 2010. 629:141-58...viral high-content screen. Moffat J, Grueneberg DA, Yang X, Kim SY, Kloepfer AM, Hinkle G, Piqani B, Eisenhaure...

...function alleles. To introduce specific genomic changes, researchers use ssDNA or dsDNA repair templates...51295 pRGEB31 rice snoRNA U3 BsaI none S. pyogenes Yang 50579 pBUN6A11 OsU3 BsaI yes, activate S. pyogenes...pRGEB32 rice snoRNA U3 BsaI yes, cut S. pyogenes Hygro Yang 62201 pHSE401 AtU6-26 yes, cut S. pyogenes Hyg Chen...pRGE32 rice snoRNA U3 BsaI yes, cut S. pyogenes Hygro Yang 62202 pKSE401 AtU6-26 yes, cut S. pyogenes Kan Chen...50929 pRGE31 rice snoRNA U3 BsaI none S. pyogenes Yang 47024 pUC gRNA Shuttle Mt U6.6 inFusion none S. ...

... protein 35 ARHGEF2 Rho/Rac guanine nucleotide exchange factor 2 CCND CCND1 CCND2 CCND3 Cyclin D CDK CDK4...Phosphatidylinositol-3,4,5-trisphosphate-dependent Rac exchange factor 2 PRKA PRKAA1 PRKAA2 PRKAB1 PRKAB2 PRKAG1... RAPGEF RAPGEF1 RAPGEF2 Rap guanine nucleotide exchange factor RAS KRAS MRAS NRAS Kirsten rat sarcoma ...protein SOS SOS1 SOS2 SOS Ras/Rho guanine nucleotide exchange factor SPRED SPRED1 SPRED2 SPRED3 Sprouty related...Tuberous sclerosis VAV1 Vav guanine nucleotide exchange factor 1 YAP1 Yes associated protein 1 Return ...

...receptor for SARS-CoV-2 and SARS-CoV is hACE2, an angiotensin receptor. Another key step is the priming of ...the links and references below: ACE2 - (Human Angiotensin Converting Enzyme 2) is the host cell receptor...nucleic acids with SHERLOCK and DETECTR . Feng Zhang's lab at the Broad Institute has utilized the natural...design resources (Link opens in a new window) Find Zhang Lab Cas13a plasmids Additional protocol for using...Medicine The proximal origin of SARS-CoV-2 . BSG - Wang, K et al, 2020, BioRxiv SARS-CoV-2 invades host ...

...into cholinergic neurons. Nat Commun. 2013;4:2183. Zhang Fetal Lung Fibroblasts Cholinergic Neurons Lentiviral...into cholinergic neurons. Nat Commun. 2013;4:2183. Zhang Fibroblasts Oligodendrocyte Precursor Cells Lentiviral...ALS Patients. Cell Rep. 2016 Jan 5;14(1):115-28. Zhang Adult Dermal Fibroblasts Neurons Lentiviral Human... Cell. 2019 Jun 6. Buganim Glial Cells Retinal Ganglion Cells AAV/CRISPR Mouse Glia-to-Neuron Conversion...Disease in Mice. Cell. 2020 Apr 30;181(3):590-603.e16. Yang iPSCs Bipolar Neurons Lentiviral Human Rapid neurogenesis...

...targeting. Cermak T, Doyle EL, Christian M, Wang L, Zhang Y, Schmidt C, Baller JA, Somia NV, Bogdanove.... A third kit, a "TALe Toolbox", from Dr. Feng Zhang’s group at the Broad Institute, will be available...available from Addgene before the end of the year. Dr. Zhang ‘s lab published a paper on TAL effectors in Nature...

...resistance from EFS promoter. Lentiviral backbone. Zhang Cas9 and Accessories for Activating Gene Expression...none Expresses dCAS9-VP64 activator with 2A GFP Zhang 61425 lenti dCAS-VP64_Blast dCAS9 (D10A, N863A) ...marker (EF1a-NLS-dCas9(N863)-VP64-2A-Blast-WPRE) Zhang 61426 lenti MS2-P65-HSF1_Hygro MS2-P65-HSF1 Hygromycin...resistance marker (EF1a-MS2-p65-HSF1-2A-Hygro-WPRE) Zhang Pooled Barcoding Libraries ID Name Description PI...

...EGFP Carol Otey 31923 pPaxillin-PSmOrange Focal Adhesions paxillin PSmOrange Vladislav Verkhusha 79999 pZyxin-miRFP703...pEGFP-C1-Fibrillarin Nucleolus Fibrillarin EGFP Sui Huang 36207 pmTurquoise2-H2A Nucleus H2A mTurquoise2 Dorus...Clathrin-coated vesicles Clathrin light chain GFP Bo Huang 54040 mEmerald-Clathrin-15 Clathrin-coated vesicles...vesicles Clathrin, light polypeptide EYFP Xiaowei Zhuang 14437 mRFP-Rab5 Early endosomes RAB5A mRFP Ari ...

...accomplish this they have taken advantage of the Zhang lab all-in-one (guide/Cas9/reporter) pX458 plasmid...desalted oligonucleotides with the appropriate overhangs: The plasmids listed here are provided as empty...Information Cloning Protocols Single guide cloning: Zhang Lab General Cloning Protocol (PDF, 237 KB) Dual ...

...through base excision repair, creating a C to T change (or a G to A on the opposite strand). Likewise,...guanosine by the cell, creating an A to G (or T to C) change. Adenine DNA deaminases do not exist in nature,...targeted AID-mediated mutagenesis (TAM) from the Chang lab and CRISPR-X from the Bassik lab , in which ...

...activator system. Cheng AW, Wang H, Yang H, Shi L, Katz Y, Theunissen TW, Rangarajan S, Shivalila CS, Dadon...

...Additionally, these viruses are able to target a broad range of dividing and non-dividing cell types with almost...Gateway cassette; See article for additional plasmids Zhang 50957 RedTrackCMV Shuttle For production of mRFP-trackable...tag; See plasmid 62622 for plasmid without Venus Wang Adenoviral Kits Plasmids containing modular inserts...

...affinity chromatography, antibodies undergo a buffer exchange using centrifugal columns. The final formulation...the protein size is confirmed. Residual Plasmid Sanger Sequencing Purified recombinant antibody preps ... The PCR product is purified, sequenced via the Sanger method, and aligned with the parental plasmid sequence...