...for one well in 24-well plate. A good example/reference is this Cell paper: http://www.cell.com/abstract... 10uM oligo stock for each well on 24-well plate, co-deliver into the cells with the Cas9/sgRNAs vector... vector, for a one well in a 24-well plate. A good reference is this paper in Cell . Click here for a ...find both single-allelic and bi-allelic cells. Single-allelic cells usually make up the majority in culture...specificity. A good reference is this Cell paper: http://www.cell.com/abstract/S0092-8674(13)01015-5 As for...allows you to sort for GFP-positive cells and to enrich for those cells that were positively transfected....CRISPR is used to engineer a locus, isolate single cell colonies, then genotype them, it is possible to ...

...channelrhodopsin-2 driven by PV+ basket cell-targeting enhancer Gordon Fishell Viviana Gradinaru 213940-PHPeB ...expressing dTomato driven by PV+ basket cell-targeting enhancer Gordon Fishell Viviana Gradinaru 213937-PHPeB ...channelrhodopsin-2 driven by chandelier cell-targeting enhancer Gordon Fishell Viviana Gradinaru 213936-PHPeB ...expressing dTomato driven by chandelier cell-targeting enhancer Gordon Fishell Viviana Gradinaru 213915-PHPeB ...Collection The Armamentarium for Precision Brain Cell Access is developing a collection of molecular genetic... reagents to gain access to many different brain cell types. The expanded collection of Armamentarium ...10aa-H2B-WPRE3-BGHpA (Alias: CN4496) Expression of SYFP2 (yellow fluorescent protein) plus separate nuclear FLAG...

...Control Boyden 83900 pAAV-mDlx-GFP-Fishell-1 Dlx GFP Control Fishell 99130 pAAV-mDlx-NLS-mRuby2 Dlx mRuby2...Control Fishell 213829 pAAV_BiCHATe27_dTomato_nlsdTomato BiCHATe27 dTomato/nlsdTomato Control Fishell 213855...Control Fishell 213936 pAAV_BiPVe4_dTomato_nlsdTomato BiPVe4 dTomato/nlsdTomato Control Fishell 213940 ...Control Fishell 213944 pAAV_BiSSTe4_dTomato_nlsdTomato BiSSTe4 dTomato/nlsdTomato Control Fishell Chemogenetics...Activator ChR2 Fishell 213830 pAAV_BiCHATe27_ChR2_mCherry BiCHATe27 Activator ChR2 Fishell 213856 pAAV_BiVIPe4...Activator ChR2 Fishell 213915 pAAV_BiLAMP5e3_ChR2_mCherry BiLAMP5e3 Activator ChR2 Fishell 213937 pAAV_BiPVe4... Activator ChR2 Fishell 213941 pAAV_BiPVe3_ChR2_mCherry BiPVe3 Activator ChR2 Fishell 213945 pAAV_BiSSTe4...

...PRKCG MTH Spinocerebellar ataxia 30 Frederic Mushinski 8418 pLTR PKC gamma PRKCG Spinocerebellar ataxia ...GFP CMV Spinocerebellar ataxia 27 Tobias Meyer 21204 GFP-N2-PKCgamma PRKCG GFP CMV Spinocerebellar ataxia...GFP CMV Spinocerebellar ataxia 25 Tobias Meyer 21215 GFP-C2gamma PRKCG GFP CMV Spinocerebellar ataxia ...CMV Spinocerebellar ataxia 23 Bernard Weinstein 21237 PKC gamma dNPS PRKCG HA CMV Spinocerebellar ataxia... CMV Spinocerebellar ataxia 21 Bernard Weinstein 21239 PKC gamma DN PRKCG HA CMV Spinocerebellar ataxia...OPTN His CMV ALS Jon Ashwell 23051 pDEST26-hOPTN-D474N OPTN His CMV ALS Jon Ashwell 23052 pDEST15-hOPTN...hOPTN OPTN GST T7 ALS Jon Ashwell 23053 pDEST17-hOPTN OPTN His T7 ALS Jon Ashwell 23054 pDEST15-hOPTN-D474N...

...genomic loci in living human cells by an optimized CRISPR/Cas system. Cell . 155(7):1479-91. PMID: 24360272... for Mapping Multiprotein Complexes in Human Cells. Cell Rep . 13(3):621-33. PMID: 26456817 Fu Y, Sander...Sontheimer EJ, Maxwell KL, Davidson AR. 2016. Naturally occurring off-switches for CRISPR-Cas9. Cell . 167(7)...described CRISPR-based genome editing in mammalian cell culture. Five years later, PubMed lists more than...orthogonality, and multiplexibility in various species, as well as the development of new applications. The first...multiple loci can be easily expressed in the same cell or organism. The ability to precisely target CRISPR...Current libraries are available for gene knockout, as well as transcriptional activation or repression. Combined...

...information Mouse and Cell Line Repositories (Link opens in a new window) Coriell Institute - cell line repository...Unpublished Joost Gribnau Human Cell Line Models Induced pluripotent stem cells (iPSCs) are a powerful system...opens in a new window) LUHMES cell line , an immortalized pre-neuronal cell line that can be differentiated... RSRT cell line collection (186.8 KB) . To order, please email [email protected] The following cell lines ...Collection and Resource Center MECP2 Animal Models Human Cell Lines CRISPR Tools Plasmids Resources The (Link ...X inactivation results in approximately half of cells expressing wild-type MECP2 and half expressing mutant...treat Rett syndrome and restore functional MECP2 in cells. Search (Link opens in a new window) JAX Mice for...

...mCherry-P2A-Cre Cre CMV RABV Jessell 73474 RabV CVS-N2c(deltaG)-Cre Cre CMV RABV Jessell 73717 pMLS328 Cre eft...Mammalian Heller 112615 pEHC Emerald and Cre-ERT2 with MCS for inserting promoter none Mammalian Heller 112616...promoter none Mammalian Heller 112618 pCAG-VHC Venus, Cre-ERT2 CAG Mammalian Heller 112619 pCAG-EHC Emerald...Cre-ERT2 CAG Mammalian Heller 112620 pCAG-mTHC TFP, Cre-ERT2 CAG Mammalian Heller 112621 pCAG-tdTHC tdTomato...allowing for direct comparison of Cre+ and Cre- cells. While Cre-lox recombination is a very useful tool...promoter like CAG, or expressed only in a subset of cells under a more specific promoter (e.g. Rho-Cre is ...promoters. Expression of both N and CCre in the same cell type allows for recombination of LoxP flanked DNA...

...to target a variety of cell populations in different brain regions. Labeled cell populations were tested...WPRE3-BGHpA AiP1379 AiE2150m SYFP2 Purkinje cells Cerebellum 208141 pAAV-AiE0390m_3xC2-minBG-SYFP2-WPRE3...regulatory sequences — termed enhancers — to drive cell type-specific expression. The Allen Institute for...differential chromatin accessibility across different cell types and cloned into an AAV backbone. Most enhancers...mouse brains. This collection contains plasmids for cell populations in the following regions: Isocortex ...Cortical subplate Striatum Pallidum Hypothalamus Cerebellum Whole brain Addgene ID Plasmid Allen Institute...-WPRE3-BGHpA AiP12320 AiE0475m SYFP2 Chandelier cells (Pvalb) Isocortex 230384* pAAV-AiE0475m-minBG-iCre...

...carrying version 1 of the CellTag barcoding library. Version 1 of the Celltag library contains 19973 barcodes...barcodes to combinatorially index cells for single-cell analysis of clonal dynamics. This library expresses...carrying version 2 of the CellTag barcoding library. Version 2 of the Celltag library contains 4934 barcodes...carrying version 3 of the CellTag barcoding library. Version 3 of the Celltag library contains 5737 barcodes...knockout pooled libraries 73178 Human sgRNA library Brunello in lentiGuide-Puro Human sgRNA library in backbone...

...Sauka-Spengler Lab Lineage Tracing Cell lineage tracing in multicellular systems typically relies on genetic... and track clones of cells over time. Examples of useful tools for studying cells that share clonal origin...Shendure Lab Simultaneous single-cell profiling of lineages and cell types in the vertebrate brain - Alex...Mumm Lab. NTR 2.0/MTZ-based cell ablation system that allows sustained cell loss and ablation of ...below: UFlip CRISPR targeted integration vectors for cell-type specific conditional gene inactivation - Jeffrey...Pantazis Lab Multibow: digital spectral barcodes for cell tracing - Sean Megason Lab Zebrabow - Albert Pan...aggregates in real time. Live imaging of astroctyes - Kelly Monk Lab. Live imaging tools to study the development...

... preparations are added to cells in culture. Three to five days later, cell cultures are inspected for... a new vial of early passage cells. Approximately 2 weeks post-thaw, cell culture supernatant is tested...involves transducing cells with serial dilutions of the lentiviral vector, harvesting cells several days later...see maps section for images). Figure 2: AAV Pro cells were transduced with either pAAV-Ef1a-mCherry-IRES-Cre... (55634-AAVrg) alone at 1.7E6 viral genomes (vg)/cell, pAAV-CAG-FLEX-rc [Jaws-KGC-GFP-ER2] (Addgene 84445...Production Lentiviral preparations are produced in 293T cells using the 2nd generation psPAX2 and pMD2.G packaging...packaging system developed by the Trono lab . Cell culture medium (OptiPro SFM) containing lentivirus is ...

...effectors to visualize subcellular positioning of target DNA sequences in living cells (TALE-mediated Genome...serial ligation. Validated in zebrafish somatic cells. Keith Joung Zhang Lab TALE Toolbox PCR/Golden Gate...ligation-independent cloning (LIC). Validated in human cells. Veit Hornung Musunuru/Cowan Lab TALEN Kit Kit comprised...pC-GoldyTALEN Golden Gate TALEN 2.0 36033 pTAL5-BB Tom Ellis Plasmids pTAL5-BB and pTAL6-BB are designed as alternative..., both in microinjected embryos and transfected cells. TALEN expression is driven by the strong CAG promoter...destination vectors and a reporter vector for mammalian cell-based validation assay that are designed for use... the evaluation of TALEN activities in cultured cells. Golden Gate TALEN 2.0 47388 pcDNA3.1-GoldenGate...

...protein-protein interactions and nerve cell function within the cell. Parkinson's Disease Parkinson’s disease...inducible vector . Harley et al. Cell Rep. 2023 Dec 26. Induce CNS cells with concomitant expression of ...with the disease and implicate a wide range of cellular processes. The vast majority of cases (90-95%)...that causes the progressive degeneration of nerve cells in the brain. Symptoms include changes in behavior...emotion, uncontrolled movements, memory deficits, as well as difficulty speaking and swallowing. HD is caused...FRET-based reporter to study tau seeding in mammalian cells. Lathuiliere et al. Mol Neurodegener. 2023 Aug 8...system, peripheral nervous system, vesicular brain cells and more. New and Noteworthy: AAV.rTH.PI.Cre.SV40...

...Drosophila cells. N terminal tagging in Drosophila cells 3.2 MB C terminal tagging in Drosophila cells 3.3 MB...following article: Natsume, et al. Cell Reports 2016 Förstemann Drosophila Cell Tagging System The Förstemann... Allen Institute for Cell Science Plasmid Collection The Allen Institute for Cell Science has produced...tagged, human, induced pluripotent stem cell (hiPSC) lines. These cell lines were created using CRISPR and...methods. Several different tagging techniques, as well as the plasmids and protocols needed for using them...tagging transcription factors with FLAG in human cell lines. To repeat the tagging in your system, use...guide the Cas9 protein to your target sequence, as well as design and clone in the homology arms for the...

...hard-to-transfect mammalian cells. In addition, "viral pseudotyping" can be used to target specific cells or tissues. ...genetic material into a cell, and even deliver this genetic material to specific cells in an organism. Figure...Genome Packaging Capacity Virus Size (nm) Cells Infected Target Cell Genome Integration Immune Response Lentivirus...because of their ability to transduce non-dividing cells. These viruses enable long-term stable gene expression...ability to preferentially infect and kill cancer cells. Adenoviruses have high transduction efficiency ... though this infectivity is limited to dividing cells. Retroviruses integrate permanently into the host...for the delivery of this genetic information to cells. Adapting plasmid DNA for use with virus-mediated...

... of the viral spike (S) protein to cellular receptors. The cellular receptor for SARS-CoV-2 and SARS-CoV...SARS-CoV-2 . TMPRSS2 - Hoffmann M, et al, 2020, Cell SARS-CoV-2 Cell Entry Depends on ACE2 and TMPRSS2 and Is...key step is the priming of the S protein by host cell proteases. The S protein of SARS-CoV-2 is primed...Human Angiotensin Converting Enzyme 2) is the host cell receptor mediating the entry of SARS-CoV and SARS-CoV...2 S protein and is involved in virus entry into cells. ( 2 ) FURIN - an enzyme that cleaves precursor ...2 S protein and is involved in virus entry into cells. ( 4 ) Cathepsin L - a cysteine endosomal protease...increasing viral infectivity.( 6 ) PCP4 - (Purkinje cell protein 4) is involved in beating of airway cilia...

...genome editing in mammalian cells (Cong et al ., Science 2013, Ran et al ., Cell 2013, Ran et al ., Nature...Matoba S, Zhang Y, Zhang F. Cell . 2013 Sep 12;154(6):1380-9. doi: 10.1016/j.cell.2013.08.021. Epub 2013 Aug...Feng G, Sharp PA, Zhang F. Cell . 2014 Oct 9;159(2):440-55. doi: 10.1016/j.cell.2014.09.014. Epub 2014 Sep... combination of CRISPR-associated (Cas) genes as well as non-coding RNA elements capable of programming...CRISPR/Cas system can be implemented in mammalian cells by co-expressing the S. pyogenes Cas9 (SpCas9) nuclease...cloning. GFP-KASH fusion facilitates FACS sorting of cells and nuclei. Return to top AAV - Cas9 mouse CRISPR-Cas9...constitutively expressing Cas9 knockin mice (Platt et al ., Cell 2014). In these mice the CRISPR-Cas9 system can ...

...-GFP-Fishell-1 Dlx GFP Control Fishell 83895 pAAV-hDlx-Flex-GFP-Fishell_6 Dlx GFP Control Fishell 83894...pAAV-hDlx-GiDREADD-dTomato-Fishell-5 hDlx Inhibitor DREADD Fishell 83897 pAAV-hDlx-GqDREADD-dTomato-Fishell-4 hDlx Activator...83894 pAAV-hDlx-Flex-dTomato-Fishell_7 Dlx dTomato Control Fishell 104055 pAAV-CAG-eYFP CAG EYFP Control...83898 pAAV-mDlx-ChR2-mCherry-Fishell-3 mDIx Activator Optogenetics Fishell 65014 pAAV-hsyn-Jaws-KGC-GFP-ER2... of Cre, and expresses NLS-EGFP in Cre-positive cells. Control Harvey 137164 pAAV-nEF-Con/Fon/Von eYFP...Cre-dependent Calcium sensor Ting 83899 pAAV-mDlx-GCaMP6f-Fishell-2 mDlx GCaMP6f expression under the control of ...of the mDlx enhancer element Calcium sensor Fishell 51084 AAV-hSyn1-GCaMP6s-P2A-nls-dTomato Syn GCaMP6s...

... in well-defined cellular sub-populations has generally been limited to single-component cellular definitions...to enable three-feature cell targeting. C) Example of viral targeting of cells expressing Cre AND Flp,... Motifs. Cell Rep. 28(13):3450-3461. PubMed (Link opens in a new window) Mingote S, Amsellem A, Kempf ... powerful molecular reagents for biology have propelled our understanding of neural circuit functionality...parameters. Figure 1: Examples of intersectional cell population definitions using recombinases. A) Cre...adeno-associated virus (AAV)-borne payloads to be expressed in cells based on a doubly-specified combination of genetic...multiple recombinase-expression strategies to define cellular sub-populations of interest, including dual-transgenic...

...not secreted, so cells must be lysed or the substrate must be delivered intracellularly in order to measure...Expression of firefly luciferase from a cell-free mammalian cell lysate Marcel Bruchez 108542 pLenti-EF1a-Luciferase-IRES-Blast-WPRE...luciferase ORF can be found in a table below, as well as plasmids that already contain a region of interest... viral infection efficiency, to study changes in cell physiology, or as a reference level of expression...ATP-independent luciferases derived from teLuc that emit yellow/green light and red light, respectively, for increased...interactions , protein dimerization , endosomal escape of cell-penetrating peptides , and more. nLuc/cLuc for split...Resonance Energy Transfer (BRET) CalFlux VNT : An intracellular calcium sensor created by fusing NanoLuc® and...

...and display of proteins on the extracellular surface of S. cerevisiae cells His-tagged versions of pFastBac... the elements needed for replication in the host cell and are designed for a user to insert a gene of ..., or express the insert they carry in the target cell. These vectors allow you to test the function of....1-mCherry - Expresses mCherry tag in mammalian cells pBI-MCS-EGFP - Tet-inducible Find...or Fusion Protein Tags and fusion proteins are excellent tools for further understanding the function... as GFP, which allows you to visualize the cellular localization of your protein. Additionally...experiments, scientists often want to create stable cell lines in which the expression cassette of interest...

...systems were tested for functionality in mammalian cells. The lac system with the inducer isopropylthio-β-galactoside... and Bujard tested the tet system in a mammalian cell system (HeLa) and found that the tet system was ... interest rtTA or TetR expression plasmid Stable cell lines can be made that continuously express a system...derivatives can be viewed in Krueger et al, 2004. Some cell culture sera may contain tetracycline or its derivatives...additional tookit plasmids third-generation rtTA On Elledge 19407 pTREtight2 Promoter contains a modified TRE...of gene expression at a single copy in mammalian cells; Expresses firefly luciferase hairpin and GFP under... under pTREtight promoter None Either Elledge 35625 pAAV-Ptet-RFP-shR-rtTA AAV; shRNA cloning vector; ...

...Amber N1 Yellow Mammalian Express a gene of interest fused to the N-terminus of Amber Amber C1 Yellow Mammalian...Venus pCEP4YPet-MAMM Yellow Mammalian Expresses mammalian optimized YPet pYPet-His Yellow Bacterial Expresses...protein interactions or conformation changes, as well as FRET standards. Plasmid...Protein Resources: Empty Backbones Biosensors Subcellular localization Optogenetics Background Förster ... Constructs to target mTurquoise2 to various subcellular compartments pCEP4CyPet-MAMM Cyan Mammalian Expresses...interest fused to the C-terminus of Amber mVenus N1 Yellow Mammalian Express a gene of interest fused to the... the N-terminus of monomeric Venus mVenus C1 Yellow Mammalian Express a gene of interest fused to the ...

...experiment, target cells are treated with the pooled library to create a population of mutant cells that are then...quantities of cells and therefore, may not be suitable for use when the number of cells is limited (e....ribosomal, cell cycle) 51043 — 51048 Knockout Human Sabatini and Lander 3rd 10 Varies FNLCR CRISPRa Cell Surface...e.g. primary cells). Most CRISPR libraries require the use of lentivirus to deliver the gRNA/Cas9 to target... target cells. Therefore, you must be equipped and authorized to use lentivirus in your lab. Once you ...organism, and the library is only compatible with cells derived from that organism. All of our current pooled...Bradley 3rd 8-10 pgRNAs 1,718 Broad GPP genome-wide Brunello 73179 (1 plasmid) 73178 (2 plasmid) Knockout Human...

...knocking down a gene can tell you much about its function and/or alter cellular activity in a way that’...also use bacteria like E. coli and their relatively well understood biology to answer many interesting questions...expression and activity thereby limiting the use of cellular resources required for the production of large...tags whose localization you can monitor in live cells (beware of artifacts!), photoactivatable tags for... tags Various Plasmids Blue/UV , Cyan , Green , Yellow , Orange , Red , Far-Red , Near-Infrared , Large... (CyPet) optimized for FRET measurements with a yellow fluorescent protein variant (YPet) found in pYPet-His...inducible expression of an N-terminally His-tagged yellow fluorescent protein variant (YPet) optimized for...

...vectors designed to express nucleases in different cell types. Zinc finger arrays can be transferred in-...vectors designed to express nucleases in different cell types. Zinc finger arrays created using ZFC reagents...nuclease domains: pST1374 (for expression in mammalian cells or zebrafish) pDW1775 (for expression in plants)... Miller et al., Nat Biotech 2007 ) in mammalian cells or zebrafish (can be used as a substitute for pMLM335... Miller et al., Nat Biotech 2007 ) in mammalian cells or zebrafish (can be used as a substitute for pMLM336... Miller et al., Nat Biotech 2007 ) in mammalian cells or zebrafish). pMLM802 (for expression of a ZFN ... Miller et al., Nat Biotech 2007 ) in mammalian cells or zebrafish). pMG10 (a '“2-in-1” ZFN cassette for...

...Laplante M, Sabatini DM. Cell. 2012 Apr 13;149(2):274-93. doi: 10.1016/j.cell.2012.03.017. PubMed PMID... (mTOR) is a key metabolic regulator controlling cell growth and proliferation. mTOR, a serine/threonine...signals to determine the nutritional state of the cell and regulates the switch from anabolic to catabolic...growth factors, including insulin. mTORC2 influences cell survival and cytoskeletal organization. Cancer Impact... promotes the conditions needed for uncontrolled cell growth and proliferation, including an abundant ...abundant supply of macromolecules and energy, as well as the presence of both pro-proliferative and anti-apoptotic...appears to signal upstream of Akt, also promotes cell survival and proliferation. mTOR Pathway Plasmids...

...adipose tissue-derived mesenchymal stem cells into pluripotent stem cells by a mutant adeno-associated viral..., 20 mM Tris at pH 8.0) Suitable for in vitro as well as in vivo delivery where the vector is diluted ...serotype also demonstrates enhanced transduction of cells in culture for ex-vivo conditioning. The AAV2(trpYF...displays enhanced transduction of hematopoietic stem cells and microglia. The AAV6(dbY-F+T-V) serotype was ...AAV-mediated transduction and targeting of retinal bipolar cells with improved mGluR6 promoters in rodents and primates...human gene therapy of glaucoma and their reversed cell entry characteristics. Gene Ther . 2020 Apr;27(3...delivery of a novel AAV vector targets ON bipolar cells and restores visual function in a mouse model of...

...molecular switches that transmit signals within the cell. GTPases have an active (GTP) and inactive (GDP)...prominent roles in signaling pathways regulating cell migration, survival, adhesion, growth, and differentiation...Cancer Impact Ras is ubiquitously expressed in all cells and all tissues. Ras can activate a variety of ...pathways, which ultimately turn on genes involved in cell growth, differentiation and survival. Ras is essential...essential for regulating the growth of the cell, however when mutations occur in the Ras gene it can be... active form, constantly activating pathways for cell growth and potentially leading to cancer. Constitutively...E2F2 E2F3 E2F transcription factor ECT2 Epithelial cell transforming 2 EGFR Epidermal growth factor receptor...

...p53 are in promoting cell cycle arrest, apoptosis, or senescence in damaged cells. The p53 name arose ...top References Unravelling mechanisms of p53-mediated tumour suppression. Bieging KT, Mello SS, Attardi ...transcription factor and tumor suppressor activated by cellular stress. Cancer...transcription factor and tumor suppressor activated by cellular stresses like DNA damage, oxidative stress, and...or E2 Cyclin G CCNG1 CCNG2 Cyclin G1 or G2 CDC25 Cell division cycle 25C Cdk4/6 CDK4 CDK6 Cyclin-dependent...member 10b E2F-1 E2F transcription factor 1 Fas Fas cell surface death receptor Gadd45 GADD45A GADD45B GADD45G...ML, Ogryzko V, Gardner K, Giordano A, Levine AS, Kelly K. Nature. 1997 Jun 27;89(7):1175-84. PubMed PMID...