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We narrowed to 784 results for: SCR;

Showing: 1 - 50 of 784 results

  1. The Open Repository of CRISPR Screens: CRISPR Screen Data in One Place

    Type
    Blog Post
    ... and the screen results. Screenshots updated 2024. Importantly, metadata for each screen is captured...Repository of CRISPR Screens. Imagine you’ve just discovered that your favorite gene was described in a CRISPR ...CRISPR screen data that might be able to help you out. What is the Open Repository of CRISPR Screens? The...of CRISPR-based phenotypic screen datasets. Original published CRISPR screen data is curated and deposited...BioGRID Open Repository of CRISPR Screens (ORCS) homepage. Screenshot updated 2024.   Using the Open...Repository of CRISPR Screens BioGRID ORCS currently houses over 890 CRISPR screens in 629 cell lines, each...cancer" returns 40 screens performed in 26 different breast cancer cell lines. Screenshot updated 2024. ...

  2. Plasmids 101: Screens vs. Selections

    Type
    Blog Post
    ...right for you. Screen variations Forward and reverse screens Forward and reverse screens involve manipulating... suppressor screens These types of screens are very similar to forward and reverse screens, except that...approaches. Screens and selections The common goals The confusion in differentiating between a screen and a ...reverse screen – all known genes are assayed for their role in a particular capacity. Drug screens, on the... time screening hits that aren’t “real” or novel for their purposes… or worse yet have a screen that doesn...phenotype. These screens employ the same downstream manipulations as a forward or reverse screen (CRISPR, drug...component. Alternative screening techniques While CRISPR and drugs dominate the screening scene, there are ...

  3. The AAVantages of AAV in CRISPR Screens

    Type
    Blog Post
    ...variety of screens, including interference, activation, and knockout. Most CRISPR screens rely on lentiviral..., AAV vector screens may be AAVantageous!       In vivo, in vitro, ex vivo CRISPR screens can be carried... For in vivo screens, mice are directly infected with the CRISPR library. In vivo screens can become costly...CRISPR Delivery Genome-wide Screening Using CRISPR Performing In Vivo CRISPR Screens Using the FITS Approach...Forward genetics screens are a valuable part of the molecular biology toolbox to identify new target ...understand the intricacies of molecular pathways. These screens have gotten larger, easier, and more comprehensive...CRISPR-based technologies over the past decade. CRISPR screens are easily customizable and enable the targeting...

  4. Tips for Improving Your Next Manuscript

    Type
    Blog Post
    ...improve your next manuscript 1. Be consistent with your figures The figures of a manuscript are the result...introduce the reader to the manuscript and engage them in the material. So the manuscript should be able to be... When Publishing a Scientific Manuscript Writing Scientific Manuscripts: Literature Searching, Reading... a publication. There is an art to composing manuscripts and the ASM Scientific Writing and Publishing...the do’s and don’ts of that week’s piece of a manuscript. The webinars were my favorite part due to the...consisted of reading that class’s section of a manuscript and critiquing it, were not too laborious, but...helped build my understanding of how to generate a manuscript. At the end of the online course, we were given...

  5. Genome-wide Screening Using CRISPR

    Type
    Blog Post
    ...genome-wide screens? The goal of a genome-wide screening experiment is to generate and screen a population...the steps involved in a CRISPR screen? Performing a forward genetic screen using CRISPR libraries is a multi-step...sorting and screened for a particular phenotype. For example, mutant cells can be used in drug screens to identify...means to carry out so-called “forward genetic screening experiments”, it is certainly the most robust....libraries are being used to perform genome-wide screens and highlight some of the reagents that have been... different genomic target. The goal of a CRISPR screen is to use Cas9 and a pool of gRNAs to identify ...CRISPR can be readily scaled up for genome-wide screening due to the broad range of potential target sequences...

  6. Plasmids 101: Blue-white Screening

    Type
    Blog Post
    ...pUC19, and pBluescript are a few common vectors. Blue-white screening is just that – a screening process...insert (blue-white screening). Other types of screening methods Although blue-white screening is probably the... that! (To read about other screening strategies, check out our screening overview blog post.) *Note:...good, but how does this help us screen plasmids? Blue-white screening in the lab Scientists discovered...the medium to ensure transcription of the lac operon.   Tips for blue-white screening Use a good control...can talk about an even more specific method of screening your cloning reaction. Being able to select for...Note: Two Popular Vectors for Blue White Screening Available at Addgene Are: pUC18 and pUC19 Let’s begin ...

  7. Custom CRISPR Screens & the Green Listed Software

    Type
    Blog Post
    ...CRISPR knockout screens: Full genome vs. custom screens Full genome CRISPR knockout screens have been used...generating a massive amount of descriptive transcriptomics data. Custom CRISPR screens designed to target differentially... gRNAs in a screen, to ensure that each gRNA has a good chance to contribute to the screen. In the end...Secondary screens  If you have generated a list of candidate genes from a full genome screen, a smaller...genes you’d like to investigate in your next CRISPR screen. Luckily, the Green Listed software can help you...software tool used to design gRNAs for custom CRISPR screens targeting a (long or short) list of genes provided...fantastic tools for unbiased discovery. With these screens, the investigator generates a cell population where...

  8. Screening for Successful Genome Editing with Digital PCR

    Type
    Blog Post
    ...in a sample. Digital PCR assays for mutation screening Now that we have introduced how digital PCR works...the bi-allelic mutations typically desired when screening clones for complete functional knockout of your...interested in harnessing the power of digital PCR screening to validate your genome edits, this paper provides...Remember that although digital PCR is a great screening tool, it still requires target DNA sequencing ... the exact nature of your mutations. A great description of the plethora of available sequencing options...PCR-Based Method for Efficient and Highly Specific Screening of Genome Edited Cells. PLoS ONE 11, e0153901–...

  9. Pairing CombiGEM and CRISPR for Combinatorial Genetic Screening

    Type
    Blog Post
    ...multiplexed CRISPR systems for high-throughput screening are vastly useful for mapping the combinatorial...can be used to combinatorially modify the genome, screen for a particular phenotype, and quickly profile...expression of other genetic elements including transcription factors (Cheng et al., 2014) and microRNAs (.... Hart, Traver, et al. "High-Resolution CRISPR Screens Reveal Fitness Genes and Genotype-Specific Cancer...Hiroko, et al. "Genome-wide recessive genetic screening in mammalian cells with a lentiviral CRISPR-guide.... Konermann, Silvana, et al. "Genome-scale transcriptional activation by an engineered CRISPR-Cas9 complex...PMC4420636. 7. Ma, Hongming, et al. "A CRISPR-based screen identifies genes essential for West-Nile-virus-...

  10. Tips for Screening with Yeast Two Hybrid Systems

    Type
    Blog Post
    ...format of some transcriptional activators: that the DNA binding domain (DBD) and transcriptional activation...identifying prey proteins after screening. It is also possible to screen libraries of bait and prey against...your rate of interactions when screening! Tips for two-hybrid screening Minimizing false positives: Run...cerevisiae in 1989 and are still used extensively to screen for molecular interactions in the cell, including...flurry of discovery in the field of eukaryotic transcriptional activation and cell biology. During this period... regardless of whether it interacts with a transcriptional activator (1a) or a repressor (1b). Scientists... these discoveries to create a new system for screening protein-protein interactions. In later years, ...

  11. Dos & Don'ts When Publishing a Scientific Manuscript

    Type
    Blog Post
    ...help the editor handling your manuscript decide on whether your manuscript fits the scope and the interest...proofread your manuscript and help you correct any language- and spelling mistakes. A manuscript that is well...journal. Publishing via open access means that your manuscript will be available to anyone who wishes to read...read it, without them needing a subscription to the journal. An increasing number of funding bodies insist...style and the things you need to include in your manuscript. What information do I need to include in my ...in the Materials and methods section of their manuscript to make resources easier to identify, find back...back, and share. Depending on the type of manuscript you are writing, it may be useful to consult the ...

  12. Plasmids 101: Screening Strategies Used in Plasmid Cloning

    Type
    Blog Post
    ... your selection, then screen Why add a screen to your cloning strategy? A screen will help you more easily...a look at a few screening methods. Blue-white screen A widely used method of screening is the blue-white...In many cases, you will need a screening strategy as well. In a screen, you aren’t killing off a portion...blue-white screening blog post for more information.   Figure 1: Result of a blue-white screen. Image...that contain your desired insert? This is where a  screen comes in. Let’s take a look at All_in_one_CRISPR...vector contains the gRNA insert or not. A blue-white screen, however, can give you this information (more on...blue-white screen, which relies on the lacZ gene. lacZ encodes the enzyme 𝛽-galactosidase which can hydrolyse...

  13. Plasmid Grant to Aid Cervical Cancer Screening in Ghana

    Type
    Blog Post
    ...Cervical cancer screening in Ghana Gladys is currently working to enhance cervical cancer screening in Ghana....cervical cancer screening for a limited group of Ghanaian women by including PCR screening for HR HPV alongside...as positive controls in her PCR screens. She hopes that these screens will both identify individuals who...to be at high risk by these screens can then undergo more routine screening allowing for early cervical...positive controls in PCR-based cervical cancer screens. This third grant represents the continuation of...tests are commonly used in Ghana, there is little screening for DNA markers like the presence of high risk... cancer screening...

  14. Performing In Vivo CRISPR Screens Using the FITS Approach

    Type
    Blog Post
    ... vivo screens in naïve T cells. More broadly, this system can be used to perform in vivo screens in other...power of in vivo screens. Figure 1: Using FITS to perform in vivo CRISPR screens to assess T cell...FITS In vitro CRISPR screens in immune cells have identified many putative regulators of immunity. However...However, in vitro screens cannot faithfully model the complex spatial, cellular, metabolic, and cytokine...vivo. Over the past five years, in vivo CRISPR screens have been used to interrogate T cells, an important...complex and physiological microenvironments. These screens have primarily been performed in activated T cells...naïve T cells. The Framework for In vivo T cell Screens (FITS) approach (Milling and Markson et al., 2024...

  15. Writing Scientific Manuscripts: Literature Searching, Reading, & Organizing

    Type
    Blog Post
    ...Beginner’s Guide to Writing Scientific Manuscripts. Any scientific manuscript will require numerous other references...Scientific Manuscripts by Johnna Roose Dos & Don'ts When Publishing a Scientific Manuscript by Maaike...in ‘writing-mode’ for a grant or a scientific manuscript, the reading will likely be more intense, but...need to be on a network with an institutional subscription), archived on PubMed Central, or in preprint...down your own results; are they the same as the description the authors give? Ask yourself if the results...filled to the brim with reprints and papers with scribbled notations in the margins. If you prefer, printing...of the Beginner’s Guide to Writing Scientific Manuscripts series on Johnna's blog. Thank you to our guest...

  16. Cas13d: Small RNA-targeting CRISPR enzymes for transcriptome engineering

    Type
    Blog Post
    ...New work from Konermann et al. and Yan et al. describes new Cas13d enzymes that average only 2.8 kb in...type VI-D enzymes are the latest tools in the transcriptome engineering toolbox. How were Cas13d enzymes...Cas13d, both groups used very general bioinformatic screens that looked for a CRISPR repeat array near a putative...Like other Cas13 enzymes, the Cas13d orthologs described in these papers can independently process their... high targeting efficiency, both of which are described below. Yan et al.: WYL-domain-containing accessory...small hairpin RNA interference, dCas9-mediated transcriptional inhibition (CRISPRi), and Cas13a/Cas13b RNA...only time will tell how large the genome and transcriptome engineering toolbox will be. It is, however,...

  17. Lentiviral CRISPR Libraries Enable Genome-Scale, Knockout Screening

    Type
    Blog Post
    ...Two new human lentiviral CRISPR library systems described in companion papers in Science in December were...Additionally, a concurrent Nature Biotechnology article describes the development of a mouse lentiviral CRISPR library... this technology.” GeCKO library for genomic screens Using lentivirus as the delivery vehicle, Neville...to enable both negative and positive selection screening. They and their colleagues including Feng Zhang...at a time,” Wang said. “It was very difficult to screen in a pooled fashion or to go after thousands of...expression. The Yusa lab has used their library to screen for genes involved in bacterial toxin resistance... is now possible to conduct systematic genetic screens in mammalian cells,” David Sabatini, a senior author...

  18. Controlling for Off-target Effects with a New Genome-wide CRISPR Screen Design

    Type
    Blog Post
    ...biological process. These screens provide an alternative to genome-wide RNAi screens, which although highly...similar pitfalls exist for CRISPR screens and how best to design these screens to controls for flaws. Recently...CRISPR knockout screen to analyze the following unanswered questions about CRISPR screen design. Unanswered...CRISPR/Cas9 screen design Are non-targeting guides an appropriate control? Many CRISPR screens use non-targeting...library was validated via a growth screen and a ricin toxicity screen since essential and non-essential...The library performed well in both screens, with the growth screen having a 1% false discovery rate and...shRNA library screens only identified  60% of the essential genes. The ricin toxicity screen had a 10% false...

  19. Hot Plasmids June 2018 - Reverse transcriptase, nanobody, and protein-DNA interaction tools

    Type
    Blog Post
    ...expressing a novel reverse transcriptase (RT), MarathonRT. Reverse transcriptases are critical tools for ...!  MarathonRT: A highly processive reverse transcriptase Article contributed by Will Arnold  Listen ...RTs in this regard and is even able to reverse transcribe the extremely long, highly structured HCV genome...structured template!). MarathonRT also faithfully transcribes RNA to DNA with low error rates compared to other... are dependent on long, error-free, reverse transcription of RNA such as long-read cDNA sequencing and...from A. victoria the functionalized nanobodies described here can be used to target any protein fused with...unbiased discovery of proteins involved in transcriptional regulation and chromatin structure at your ...

  20. Pooled CRISPR Libraries Offer Genome-Wide Control for Large-Scale Functional Screens

    Type
    Blog Post
    ...gain-of-function screening when combined with a compact pooled sgRNA library. Our SAM-mediated screens exhibited... sgRNAs for each transcription start site in 15,977 human genes. The transcriptional repressor (CRISPRi...the next step in high-throughput, genome-wide screening,” Konermann said. “If you want to activate all...and make sure it’s working. You can only do that screen with a system you are really confident about working...Konermann and her colleagues in Feng Zhang’s lab describe just such a system, which they call CRISPR/Cas9...SAM is an engineered protein complex for the transcriptional activation of endogenous genes and it consists...The next insight was to assemble a synthetic transcription activation complex consisting of multiple distinct...

  21. CRISPR Library Amplification

    Type
    Protocol
    ...during the scraping process. Add each scrape into a 50 mL conical tube on ice pooling the scrapings of two...spreader and cold LB to scrape bioassay plates and remove bacteria. Use one scraper for all plates. Use two...mL cold LB to each plate for each scrape and use spreader to scrape plates. Pro-Tip Pushing motion is ... each tube on ice while scraping. Repeat addition of 10 mL cold LB and scrape for each plate until agar...plasmids in a single sample. They are often used for screening, barcoding, or other high throughput multiplexed...activating pulse and follow all specifications described in the equipment manual. Immediately add 1 mL ...µl aliquots of cell/DNA mixture used (8 in the described protocol) using a new vented falcon tube for every...

  22. Protocol - pLKO.1 – TRC Cloning Vector

    Type
    Protocol
    ...transforming into bacteria D. Screening for Inserts D.1 Recommended materials D.2 Screening for inserts E. Producing...10878/ . Description Vector Element U6 Human U6 promoter drives RNA Polymerase III transcription for generation...non-hairpin insert. pLKO.1 – scramble shRNA Negative control vector containing scrambled shRNA. psPAX2 Packaging...greater yield of DNA. Back to Top D. Screening for Inserts You may screen for plasmids that were successfully...generation of shRNA transcripts. cPPT Central polypurine tract, cPPT, improves transduction efficiency ...the pLKO.1 TRC-cloning vector. Addgene Plasmid Description pLKO.1 – TRC control Negative control vector ...R0101S NcoI NEB #R0193S Agarose Sigma #A9539 D.2 Screening for Inserts Day 1: 1. Inoculate 5 colonies from...

  23. Isolating a Monoclonal Cell Population by Limiting Dilution

    Type
    Protocol
    ...#52962 ) and is described in Improved vectors and genome-wide libraries for CRISPR screening. Sanjana NE,...This protocol describes how to generate a monoclonal cell line from a polyclonal pool of stable cells...Protocols Viral Service Introduction This protocol describes how to generate a monoclonal cell line from a ...type will be able to grow under the conditions described in this protocol. Conditioned medium (details ... specific conditions used. Each 96-well plate described in this protocol could, in theory, give rise to...protocol was performed at Addgene on the A549 cells described in our sample data , each 96-well plate gave rise...monoclonal lines have been sufficiently expanded, screen the lines for transgene expression and/or other...

  24. Video Library

    Type
    Protocol
    ...collection of Addgene's video content, including how-to screen captures, lab procedure protocols, and career advice...procedures. (Link opens in a new window) Video Link Description Related Page Making LB Agar Plates Create plates... from an Agarose Gel Protocol How-To Videos Screencapture walk-throughs to help guide you through Addgene's...resources. (Link opens in a new window) Video Link Description Related Page MTA Guide - Requests Your guide ...Addgenies! (Link opens in a new window) Video Link Description Jessica Welch, PhD In this first installment ...

  25. Protocols for Molecular Biology, Plasmid Cloning, and Viral Preps

    Type
    Protocol
    ...designed to help you get started in the lab. Name Description (Link opens in a new window) Link to Video Personal...blocks for many more complicated procedures. Name Description (Link opens in a new window) Link to Video Making...constructing and analyzing your plasmids. Name Description (Link opens in a new window) Link to Video Restriction...titering and testing your virus preparations. Name Description Link to Video General Transfection Introduce ...Protocols for common antibody applications. Name Description Link to Video Transfection Introduce plasmid ...

  26. Affinity Purification of Recombinant Antibodies with Protein A or Protein G

    Type
    Protocol
    ...This protocol describes how to affinity purify recombinant antibodies from cell culture supernatant using...Western Blot Protocol Introduction This protocol describes how to affinity purify recombinant antibodies ... the reservoir to a LoBind tube. Pro-Tip Do not scrape the filter with the pipette tip. Determine the ...

  27. Antibody Validation Using the Indirect ELISA Method

    Type
    Protocol
    ...This protocol describes how to run an indirect ELISA against a purified antigen to demonstrate the expected...Protocol Antibody Guide Introduction This protocol describes how to run an indirect Enzyme-Linked ImmunoSorbent...antigen to validate an antibody. This protocol will describe how to create a standard curve using known concentrations...

  28. AAV ddPCR Titration

    Type
    Protocol
    ... Biology Reference Introduction This protocol describes how to use droplet digital PCR (ddPCR) to titer...the 96-well plate sealer by gently touching the screen. Prepare the Serial Dilution Place a 48-well dilution...the plate sealer is not at temperature, touch the screen on the plate sealer to allow it to get to temperature...

  29. Lentivirus ddPCR Titration

    Type
    Protocol
    ...Guide Virus Blog Posts Introduction This protocol describes how to use droplet digital PCR (ddPCR) to titer...the 96-well plate sealer by gently touching the screen. Preparation of the Master Mix Place a ddPCR plate...the plate sealer is not at temperature, touch the screen on the plate sealer to allow it to get to temperature...

  30. Protocol - How to Create a Bacterial Glycerol Stock

    Type
    Protocol
    ...overnight culture to 500 μL of 50% glycerol in a 2 mL screw top tube or cryovial and gently mix. Notes: Make...use a sterile loop, toothpick or pipette tip to scrape some of the frozen bacteria off of the top. Do ...

  31. Water Bath Protocol

    Type
    Protocol
    ...bath on and set the appropriate temperature as described in your protocol. How to set the temperature will...water bath even if you are using disinfectants as described above in step 3. You will also need to maintain...

  32. Immunocytochemistry

    Type
    Protocol
    ...This protocol describes the basic steps for fixing and staining cells in culture with a primary antibody...antibodies to detect antigens in cells. Here we describe the basic steps for fixing and labeling cells ...

  33. Using a Light Microscope Protocol

    Type
    Protocol
    ...resolution (the ability to distinguish between two discrete objects). Figure 1 depicts an image of a compound... sure to take care of your microscope. Bumps, scratches, and dust can easily impair microscope performance...

  35. Western Blot

    Type
    Protocol
    ...This protocol describes the basic steps for lysing cells, determining total protein concentration in ...specific to the protein of interest. This protocol describes the basic steps for lysing cells, determining ...

  38. Gibson Assembly Protocol

    Type
    Protocol
    ...make your own. Transform bacteria with the DNA and screen for the correct plasmid product by restriction ...

  39. General Transfection

    Type
    Protocol
    ...Protocols Viral Service Introduction This protocol describes a general method for transfecting mammalian cells...

  43. Pouring LB Agar Plates

    Type
    Protocol
    ...maximum of ~100 colonies Pro-Tip If you will be screening a large number of colonies, we recommend using...

  46. Kit Free RNA Extraction

    Type
    Protocol
    ... Biology Reference Introduction This protocol describes how to extract RNA from tissues or cells without...

  47. Plasmids 101: Terminators and PolyA signals

    Type
    Blog Post
    ...initiation step of gene transcription; today we'll provide an overview on how transcription stops, or termination...from the transcription machinery. Terminators are found downstream of the gene to be transcribed, and typically...generally serves to promote transcript longevity in eukaryotes and promote transcript degradation in prokaryotes...thought to destabilize the transcription complex, initiating cleavage of the transcript.   Figure 1: ...expression levels. Transcription Termination Rho-dependent termination DNA transcription Polyadenylation...both prokaryotic and eukaryotic systems must be transcribed into RNA, which occurs in three phases: initiation...sequence-based element, is to define the end of a transcriptional unit (such as a gene) and initiate the process...

  48. New Optimized Genome-wide CRISPRko, CRISPRi, and CRISPRa Libraries

    Type
    Blog Post
    ...to conduct genome-wide gain of function screens. These screens can help reveal the function of lowly expressed...large-scale functional screens Read this blog post about genome-wide screening using CRISPR Resources...allowed scientists to easily perform genome-wide screens to effectively and efficiently investigate gene... per gene to increase confidence in the genetic screen. However, there are benefits to optimized libraries...resources and are particularly useful when trying to screen a limited number of cells like primary cells or... they conducted genome-wide negative selection screens with both A375 (melanoma) and AT29 (colon cancer...metrics to assess library performance in viability screens. Based on these metrics the Brunello library was...

  49. Plasmids 101: The Promoter Region – Let's Go!

    Type
    Blog Post
    ...initiating the transcription of your insert into RNA. In practice, the term "promoter" describes the combination...Promoter   Primarily used for RNA     transcript     Description Expression Additional considerations ...Primarily used for Description Expression Additional considerations T7 in vitro transcription/ general expression...for in vitro transcription, the promoter drives either the sense OR antisense transcript depending on ...controls the binding of the RNA polymerase and transcription factors, therefore promoters play a large role...

  50. New Tools Enable CRISPRa for Neuroscience Applications

    Type
    Blog Post
    ...engineered combinations of transcriptional activators to induce gene transcription at genomic sites targeted...found here. High-throughput screening strategies Although the tools described above make CRISPRa more accessible...subgenome-scale CRISPRa screening approach in embryonic stem (ES) cells to determine which transcription factors promote...this technological gap, a trio of recent papers describe the development of CRISPR activation (CRISPRa)...-optimized viral vectors, and high-throughput screening approaches. Here, we’ll highlight these recent...CRISPRa much simpler to implement in genome-wide screens over cDNA libraries. However, using CRISPRa to ...and RNA components, and several sgRNAs must be screened to identify effective ones. Simplified in vivo...
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