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We narrowed to 369 results for: Cre expression vector

Showing: 181 - 200 of 369 results

  1. What's New in CRISPR - March 2020

    Type
    Blog Post
    ...fluorescently labeled piggyBac vectors to deliver uniform and sustained expression of multiplexed gRNAs in human...CRISPRa depended on continued expression of dCas9 and gRNAs whose expression varies depending on transgene...The lab expressed both dCas9 and the ctrA sgRNA and saw that the levels of ctrA mRNA decreased after 20...demethylation Optogenetic repressors of gene expression in yeast Megan McClean’s lab generated an optogenetic...optogenetic toolkit that controls repression of gene expression in Saccharomyces cerevisiae. The system consists...the nucleus where dCas9-Mxi1 can repress gene expression. In the dark, the NLS is inaccessible and the... in Caulobacter cells for ctrA gene expression using various dCas9. Image adapted from Guzzo et al., 2020...

  2. MXS Chaining

    Type
    Blog Post
    ...structures in live cells. Four individual expression constructs were created, each containing a unique fluorescent... construct. This strategy can be used to build vectors with diverse functions, and thus, is used in many...localization, as well as cellular processes like gene expression and cell migration (Sladitschek and Neveu, 2015...enhancers, polyadenylation signals, inducible gene-expression sequences, and more. By combining these components...cell cycle dynamics, titrate inducible transgene expression, or a variety of other clever applications. ...flanked with a CMV promoter (to drive high-level expression) and a polyA signal (to terminate transcription... HeLa cells show robust labeling, with strong expression and detection of each fluorophore at the respective...

  3. AAV Q&A with Tim Miles

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    Blog Post
    ...This will be highly dependent on vector dose. The color diversity in Vector Assisted Spectral Tracing (VAST...drop in transduction coverage compared to just one vector.   Q2: Do you think it is possible to make an efficient...this scenario, you may be best served by a dual vector, split intein approach.   Q3: What’s the best method...for model organisms.    Q11: How consistent is expression is when injecting CAP-B10 or CAP-B22 in marmosets...Plasmids Adenovirus Guide Page Subscribe to Viral Vectors Blog Posts ...flexibility of AAV genome design in remarkably creative ways. The continued development of more and smaller...

  4. Five Popular Model Organisms

    Type
    Blog Post
    ...lab who created a set of modular vectors that can be used to optimally control gene expression in specific...spatiotemporal expression of a gene of interest. This type of control over gene expression is possible in... al., 2009). By injecting mice with retroviral vectors that harbored different mutations found in leukemia...that uses heat induction to control the temporal expression of different transgenes, the Stainer group determined...micro-injecting DNA into the worm, such as animals that express fluorescent markers (Mello et al., 1991). Genes...interference (RNAi) by feeding the worm bacteria expressing specific plasmids (L4440) that contain your gene...lifetime. Thus any genetic modification, such as expression of fluorescent protein or disruption of a gene...

  5. Hot Plasmids December 2018 - C. elegans RNAi, TurboID, Detecting Viral Outbreaks, and Protein Binding Specificity Tools

    Type
    Blog Post
    ...switch to be expressed. In this case, the gene encodes the LacZ𝛼 protein. Once expressed, LacZ𝛼 complexes... RNAi. Recently the Vellai lab has created an improved RNAi vector that leads to significantly stronger... RNAi is commonly achieved by feeding E. coli expressing dsRNA to C. elegans. These dsRNAs target specific...for mutant genetic backgrounds. The standard RNAi vector, L4440, places a target gene between two inverted...mutant animals that are genetically altered to overexpress endogenous RNAi pathways. These mutations however...non-specific nucleotides into the dsRNA leading to increased RNAi efficiency. Find the T444T plasmid here... used yeast display-based directed evolution to create APEX2. They have now applied the same directed ...

  6. Deep Mutational Scanning with One Pot Saturation Mutagenesis

    Type
    Blog Post
    ...codon site of interest can be cloned into an expression vector with Gibson assembly. While this is a comprehensive...relies on the generation of a plasmid library that expresses all desired variants of a protein. Applying a ...selective pressure winnows the pool down to plasmids expressing variants with optimal function. High-throughput...is generated by transforming a backbone phagemid vector into an E. coli strain that is deficient in dUTPase...2 instances where it does. 1) When the backbone vector has multiple BbvCI sites. If these sites have opposite...polymerase, 2) increased PCR extension time, 3) increased concentration of MgCl2, 4) increased concentration...PCR reaction is created by making some or all of the following modifications: 1) increased concentration...

  7. Multicolor Animals: Using Fluorescent Proteins to Understand Single Cell Behavior

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    Blog Post
    ...promoter is expressed. Cre-mediated deletion of one or several of these genes permits the expression of genes...copy of the transgene expressing either YFP or CFP, and cells that do not express Cre fluorescing with the...activate the expression of a different fluorescent protein. Therefore, each cell could express a unique combination...fluorescent protein genes, Cre recombination is required before fluorescence can occur. Cre recombinase can be...developing fish via Cre protein microinjection or crossing of Zebrabow animals to an inducible Cre line. The Schier...any cell type when crossed with mice expressing the appropriate Cre allele. The Confetti mouse contains...so that XFP are only expressed following a recombination event. Tissue-specific Cre recombination in this...

  8. Let There Be LITE Plasmids

    Type
    Blog Post
    ...those optogenetic controls, gene expression levels reliably increased in vitro by a factor of 10 to 20...functions, Konermann needed tools to manipulate the expression of specific genes in the brain in reversible ...Zhang showed they could package LITEs in viral vectors and genetically target them to specific cell populations...demonstrated use of the LITEs to control gene expression in mouse neurons and in the brains of living ...system could be by applying it to altering gene expression via epigenetics with the addition of histone ...different effectors. I’d encourage people to get creative with them for their own uses.” Konermann said ...found here.) So tell us, readers, how will you get creative with LITE? Reference: Silvana Konermann et al...

  9. Plasmids 101: Expressing shRNAs for RNA Interference

    Type
    Blog Post
    ... are transiently expressed using either AAV backbones or species-specific expression plasmids. You can...cloning strategies for constructing shRNA expression vectors. BMC Biotechnology, 6(1). https://doi.org...processing and silencing pathways. Created on BioRender.com.   Expressing shRNAs using plasmids The design...sequence. Created on BioRender.com. When it comes to the backbone, shRNAs are most often expressed constitutively...option for altering the expression of specific mRNAs. shRNAs are a quick way to create stable knockdown cells...be applied to. When it comes to altering gene expression, CRISPR technologies have gradually taken over...processing Following delivery into cells using viral vectors or plasmids, shRNAs hijack the endogenous miRNA...

  10. Single-cell tracking of lineage and identity with CellTag

    Type
    Blog Post
    ...were detected over a 10-week period. Their expression decreased over time but they were not silenced.    ...), now allows scientists to identify the gene expression patterns unique to the few cells that successfully...Unfortunately scRNA-seq only captures the gene expression of cells at a single time point, making it difficult...difficult to investigate how early expression patterns influence reprogramming outcomes. CellTag from ...single-cell level, thus linking ancestral gene expression patterns with reprogramming outcomes (Biddy et...MEFs were reprogrammed into iEPs by retroviral expression of transcription factors Foxa1 and Hnf4α, a process...lineage trees and comparing them with the gene expression changes of more than 100,000 cells undergoing...

  11. Announcing the Winners of the 2021 Michael Davidson and Roger Tsien Commemorative Conference Awards

    Type
    Blog Post
    ...working on developing new surface display vectors to improve expression and trafficking of these biosensors....gray (PAG), with red neurons expressing tdTomato and green neurons expressing cFos. Image from Elizabeth...B) CR-Cre mice demonstrate pain response due to blue light stimulation of dual virus-expressing CR neurons...verification of the dual virus expression in CR neurons of the spinal cord. Created with BioRender.com. Image... Figure 2: Micrographs of the biosensor expressed in the human cell line, Hep3B. From left to right...opioid withdrawal with tdTomato and assessed cFos expression. She will explore the connectivity patterns of...chemogenetics to study the activity of parvalbumin (PV)-expressing neurons, neurons that the lab previously found...

  12. Plasmids 101: Gene, enhancer, and promoter trapping

    Type
    Blog Post
    ...fundamentally the reporter construct monitors the expression of a nearby gene. The reporter gene serves as...as a proxy for the endogenous gene – its expression levels (high/low) and patterns (tissue/timing) can...that the gene which GFP was monitoring was being expressed in the same pattern. These systems suggest the...function of a gene. For example, they eliminate the expression of the gene (by CRISPR knockout, shRNA depletion...Tissue-specific genes – the genes of interest may not be expressed in the tissue/cell line you are working with. ...the gene as a result but will still allow for expression pattern monitoring. Enhancer traps Like promoter...trapping of endogenous enhancers to stimulate the expression of the reporter. The system is made up of a reporter...

  13. Evolution of Brainbow: Using Cre-lox for Multicolor Labeling of Neurons

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    Blog Post
    ... how Cre-lox recombination works. Advances in Cre-lox recombination have included inducible Cre, such...membrane tethered cyan). Without Cre recombination, RFP is expressed. Cre can mediate one of two deletions... construct, OFP (orange), is expressed when Cre is absent; stochastic Cre excision results in one of three...remaining cassette can invert as long as Cre remains expressed; once Cre is removed from the system, the fluorescent...Cai et al. also created an Autobow construct containing both Cre and XFPs. Cre production drives recombination...manipulate gene expression, but other gene manipulation systems remain valuable to biology. Cre-lox recombination...temporally control gene expression, especially in in vivo models, and new Cre-lox based technologies are...

  14. Common Injection Routes in Mice

    Type
    Blog Post
    ...Genome Engineering Guide CRISPR Mammalian Expression plasmids Cre-Lox plasmids References Prathipati, P.,...be injected, including cells, drugs, DNA, viral vectors, and viruses. All researchers must go through ...choice for administering different drugs and viral vectors. Like IV injections, intraperitoneal injections...for the delivery of various therapies and viral vectors. This injection sounds very disruptive, but surprisingly... This is most often used for delivery of viral vectors, though drug treatments are also possible. One ...representation of common injection routes in mice. Image created with biorender.com.   Needle techniques Intravenous... often used when injecting a cell suspension to create xenograft models. Restraint: back scruffing or ...

  15. Tips for a 1st Time CRISPR User (by a 1st Time CRISPR User)

    Type
    Blog Post
    ...cells to express Cas9. In these particular experiments, we generated A549 cells stably expressing Cas9 via...wildly variable Cas9 expression (Figure 1). This is a problem because Cas9 expression affects growth rate...variable Cas9-expression was across the different cells (Figure 3).   Figure 2: Cas9 expression in monoclonal...Checking Cas9 expression Overview: By Western blotting (Figure 2, pool), I knew the cells expressed Cas9, but...thought I could see GFP expression with a fluorescence microscope, but the GFP expression was so low (even in...against GFP expression, but I didn’t. 2. In the original publication they see GFP expression in about 20%... MOIs. 2. Create a monoclonal Cas9-expression cell line. Especially if you’re doing a screen. I made a...

  16. Hot Plasmids - October 2022

    Type
    Blog Post
    ...constructs are currently available as Cre-dependent, EF1ɑ-driven viral vectors (AAV1) - and you can also find...B) Cortical slice of HcKCR1-EYFP and tdTomato expressed layer 2/3 neurons in mouse. C) Action potential...voltage traces of action potentials in a HcKCR1-expressing mouse neuron without (top) and with (bottom) ... blot for the myc-tagged protein GFP-smFP-myc expressed from Plasmid 98926. See the Applications section...called JEDI-2P. Using a custom, high-throughput screening platform to apply saturating mutagenesis on 21... 1: Summary of the approach and outcomes of the screen for improved genetically-encoded voltage indicators... in-vivo imaging of voltage fluctuations. Image credit: Liu Z, et al. Cell. 2022.  Find JEDI-2p viral...

  17. Plasmids 101: Transformation, Transduction, Bacterial Conjugation, and Transfection

    Type
    Blog Post
    ... information on viral vectors, including transduction download our Viral Vectors 101 eBook. Bacterial ...can be used to create both transient cell lines, where a gene of interest is expressed but not integrated...bacterial or eukaryotic cells via a virus or viral vector. One example are bacteriophages that attach to ...scientists commonly use phagemids, a DNA cloning vector that contains both bacteriophage and plasmid properties...application of an electrical charge to cells, to increase cell membrane permeability and thus transformation...DNA can be accidentally packaged into the newly created phages. Transfer of this DNA to another cell is...Electroporation on the other hand uses electrical pulses to create transient pores in the cell membrane that genetic...

  18. Hot Plasmids: Summer 2025

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    Blog Post
    ...synapses. In addition, ATLAS itself could be expressed Cre-dependently to allow tracing exclusively from... mGold2. Top: TIRF-SIM imaging of COS-7 cells expressing LifeAct fused to mVenus, mGold2s, or mGold2t....but TnpB is small enough to be carried by viral vectors (~400 amino acids, less than a third of the size...thousands of eligible AAV plasmids to order viral vectors ready to use in your lab. Find Addgene’s AAV6 and...postsynaptic AMPA receptor and is endocytosed, the Cre recombinase payload activates a floxed reporter gene... 5: Mechanism of ATLASCre. VAMP2 = synaptobrevin 2; At = ALFA-tag; BACE = β-secretase; AF = AMPA.FingR...TnpB) or a gRNA (Cas9). Drawings not to scale. Created with BioRender.com. Steven Jacobsen and Jennifer...

  19. Antibodies 101: Affinity Tags

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    Blog Post
    ... single protein is expressed, with the intent of purification, it will be expressed along with all other...The MBP tag, although large, can actually increase expression and solubility of the tagged protein (Fox...other cellular proteins within the expression system (bacteria, yeast, mammalian, etc.). To individually...about that protein that no other protein in the expression system has. This is where affinity tags come ...peptides which do not naturally exist in most expression systems that can be added to the N or C terminus...on: The protein you are trying to purify The expression system If you want the tag to remain on your ...Ready to start tagging? We are here to help with vectors, protocols, antibodies, and more!   Resources ...

  20. Antibodies 101: Designing Your First Flow Panel

    Type
    Blog Post
    ...transgenic protein for the HEK 293T cells to express: CD45. Upon expression, CD45 is incorporated into the plasma...CD45, but is separately expressed. (The reason for having a separately expressed transduction reporter ...manage to express CD45. Thus, if transduction efficiency is much higher than CD45 expression, this could... example, point at issues with the double-expression vector.) In our flow experiment, we'll want to identify...CD45+ cells (CD45 expression) and assess our transduction efficiency (eGFP expression).     The first ...detected through a cell surface label. The viral vector also encodes for eGFP, which serves as a transduction...rest of our panel. Lastly, we want to detect the expression of our protein of interest: CD45. I am using ...
Showing: 181 - 200 of 369 results