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We narrowed to 849 results for: NTS

Showing: 261 - 280 of 849 results

  1. Hot Plasmids April 2018 - Protein Degradation, Nanoscopy, FIRE-Cas9, and Yeast Expression Tools

    Type
    Blog Post
    Published
    April 23, 2018, 2 p.m.
    ...deliver these components together using electroporation. In these bulk cell experiments, Clift et al. ...proteins, and it can even distinguish between protein variants when used with a variant-specific antibody. TRIM21...need to deliver the target antibody. TRIM-Away reagents and reporters are available from Addgene - we ...down the difficulty of your protein knockdown experiments! The TRIM-Away plasmids can be found here. Clift...varying which sgRNAs are used with FIRE-Cas9, experiments can be designed to target multiple loci at once...factor. They then induced cultures of these transformants with various concentrations of estradiol and...

  2. Rosella: A Fluorescent pH-Biosensor for Studying Autophagy

    Type
    Blog Post
    Published
    April 13, 2017, 2:30 p.m.
    ...Greek for “self-eating”) is induced by a lack of nutrients and targets cytosol and organelles to the vacuole...double-membraned structures containing undigested cytoplasmic contents. Autophagosomes that have fused with the vacuole...lysosome are trickier to identify because their contents can be at various stages of degradation. Additionally...Atg8p or LC3 doesn’t provide information about the contents of the autophagosome nor does it provide information...sequence. Under normal growth conditions, both variants are excluded from the vacuole and emit red and...the cytosol-targeted and mitochondria-targeted variants are available from Addgene. References 1. Rosado... L. K., Graham, T. E. (2015). Rapid parallel measurements of macroautophagy and mitophagy in mammalian...

  3. CRISPRainbow and Genome Visualization

    Type
    Blog Post
    Published
    Feb. 28, 2017, 3:30 p.m.
    ...line./ Each color represents a gRNA targeting a specific chromosome. This image represents the composite ...composite of fluorescent measurements from red, blue, and green channels overlaid on a bright-field image. Image...fluorescence from unbound GFP-dCas9. With these improvements, labeling efficiency was similar to that of ... other orthologs. Ma et al. tested their dCas9 variants using gRNAs specific for telomeric sequences and...distance! To the authors’ knowledge, this work represents the first mapping of intrachromosomal loci, a... live cells. However, they note that certain improvements could make CRISPRainbow even better. Adding ...

  4. Plasmids 101: CcdB - The Toxic Key to Efficient Cloning

    Type
    Blog Post
    Published
    Dec. 8, 2016, 3:30 p.m.
    ...specific components that are key to the cloning reaction. Understanding the specific components is essential...choosing the correct cloning method for your own experiments, and here we will focus on a unique gene that...on to find out how ccdB can make your cloning experiments a little easier. One of the most time-consuming...choose the most appropriate method for your own experiments. Please check out Addgene’s Choosing a Molecular...these resources further. Feel free to offer your comments and suggestions, and even to submit a guest blog... al. “F plasmid CcdB killer protein: ccdB gene mutants coding for non-cytotoxic proteins which retain ...

  5. Typing CRISPR Systems

    Type
    Blog Post
    Published
    March 18, 2025, 1:15 p.m.
    ...transposase (CAST) that can insert large fragments of DNA. The V-F variants, which are among the smallest Cas...on main function and composition; subtypes and variants are typically based on species of origin. There...currently two classes, six types, 33 subtypes, and 17 variants. Figure 1: Overview of the CRISPR classification...mechanisms, with the biggest differences being the components present in the Cascade complex. Figure... Cas2, which are critical for inserting short fragments of foreign DNA into the CRISPR array. Lastly, ...CRISPR–Cas systems: a burst of class 2 and derived variants. Nature Reviews Microbiology, 18(2), 67–83. https...Insights from CRISPR Loci in Integrative Conjugative Elements of Acidithiobacillia. The CRISPR Journal, 4(5)...

  6. Hot Plasmids and Viral Preps - January 2021

    Type
    Blog Post
    Published
    Jan. 5, 2021, 2:15 p.m.
    ...dimerizes bringing the C-terminal and N-terminal Cre fragments together and permitting recombinase activity. ...Biggiephage can eliminate competing mobile genetic elements. Image by Basem Al-Shayeb in the Doudna Lab. ... more compact genome editors due to delivery constraints and scientists seeking a wider range of targetable... to image over 3 million cell types expressing variants of Yellow Fluorescent Protein (YFP). The team ... As such, mGold is expected to contribute to experiments that require high photostability, such as time-lapse...compared to current N-terminal linked SaCas9 ABE variants. It's also one of the smallest AAV-deliverable...dLight1.3b here Looking for a knock-out in your experiments? AAV-flex-taCasp3-TEVp from Nirao Shah's and ...

  7. CRISPR 101: Anti-CRISPR Proteins Switch Off CRISPR-Cas Systems

    Type
    Blog Post
    Published
    July 23, 2020, 1:20 p.m.
    ... CRISPR-Cas technology is constantly evolving. Variants of Cas proteins can be used for genome editing...In this study, the labs found that anti-CRISPR prevents Cas9 activity in HEK293 by reducing editing from... AcrIIC2 interactions with Cas9 interferes and prevents the correct assembly of the crRNA-Cas complex ...binds to the HNH endonuclease domain of Cas9 and prevents target DNA cleavage (Harrington et al., 2017)....   Five ways to anti-CRISPR proteins in your experiments 1. Reduce off-target effects Prolonged Cas activity...shut off Cas activity? Using inhibitor timing experiments, the Doudna lab found that at least 50% of on-target...vectors. Having a way to shut off CRISPR-Cas systems prevents the off-target effects and cytotoxicity that result...

  8. Nanoblades: Tiny CRISPR Ninjas for Genome Editing Difficult Cells

    Type
    Blog Post
    Published
    Sept. 26, 2019, 12:50 p.m.
    ...the following four components into 293T cells. Figure 1: The key components of Nanoblades. Image... deliver Cas9 variants, such as a transcriptional activator dCas9-VPR, if these variants are fused to ...CRISPR applications. Let’s take a look at the key components of Nanoblades, how their editing efficiency and...for HDR editing and mouse-model generation. Key components of Nanoblades Nanoblades are a Cas9 RNP delivery...transgenic mice by microinjecting zygotes with CRISPR components. However, this sometimes requires intracellular...

  9. CRISPR in the Clinic

    Type
    Blog Post
    Published
    Nov. 2, 2023, 1:15 p.m.
    ...correction in adult stem cells isolated from two patients with cystic fibrosis (Schwank et al). They were...giving this treatment a CRISPR spin. CRISPR CAR-T treatments for leukemia, lymphoma, and some solid tumors...these trials has been outstanding, with almost all patients experiencing sustained and substantial relief ...are being finalized, and it is very likely the treatments will be approved for use in the next few months... a desired edit, but this particular class of treatments has quite the opposite aim: to destroy. One treatment...just don’t have the time to talk about. Other treatments currently in clinical trials include, but are...Intestinal Stem Cell Organoids of Cystic Fibrosis Patients." Cell Stem Cell. 13, 653-658 (2013). Dimitri,...

  10. High Complexity Golden Gate Assembly with NEB’s DAD

    Type
    Blog Post
    Published
    Aug. 1, 2023, 1:15 p.m.
    ...to split it into multiple fragments. You can define the number of fragments, and the “search windows” ...number of fragments, you can also ask it to give the maximum or minimum number of fragments, or defined...assembled a 40 kb T7 phage genome from 52 (!!) fragments in one very Golden (Gate) pot (Pryor et al., 2022...high-fidelity assembly for sets of up to 10-12 fragments (Pryor et al., 2020), and randomly selected non-palindromic...reaction time of 15 hours. This suggests that 52 fragments may be near the upper limit of what can be accomplished...

  11. Viral Vectors 101: Virus Safety

    Type
    Blog Post
    Published
    June 22, 2023, 1:15 p.m.
    ...One such control is dividing up the essential components of a virus into different plasmids. For a retroviral...enter (and thus infect) cells, this safety feature prevents the virus from replicating and spreading in non-target... Figure 1: Separation of retroviral packaging components onto distinct plasmids for safety.    Viral...biohazard policies. Administering Virus Many virus experiments involve needles or scalpels (sharps). Cuts and...equipment are one of the most common forms of lab accidents, so extra precaution should be used to reduce ...of an accident (the most common reason for lab accidents is not following safety protocols.) If administering...Viral Vectors 101: Types of Viruses Viral Vector Elements Tips for Preventing Viral Recombination  ...

  12. Adeno Associated Virus (AAV) for Cell and Gene Therapy

    Type
    Blog Post
    Published
    Nov. 7, 2017, 1:59 p.m.
    ...T-cells were transferred back into the infants. Almost all patients have achieved persistent immunological...To date, gene therapy has been used to treat 10 infants with this disease. To do so, their T-cells were...administration of rAAV2 carrying the human gene RPE65, patients with clinical diagnosis of LCA and mutations in... in RPE65 (n=3), showed significant improvements in vision over a period of 1 to 3 years – the treatment...Anti-AAV antibodies developed by the immune system – patients’ own immune systems can develop antibodies that...delivery tools that are under consideration, recent advancements in various AAV technologies have vastly improved...

  13. Illuminating Epigenetics with A FRET Based Biosensor

    Type
    Blog Post
    Published
    Nov. 19, 2015, 3:30 p.m.
    ...mortality determined by nutrition during parents' and grandparents' slow growth period." European Journal... histones, the more compacted it becomes. This prevents the cellular machinery from physically interacting...were experienced in a period of life when the grandparents’ gametes were developing, an epigenetic change...intellectual and developmental disability, jerky movements, and spontaneous laughter or smiling, whereas ...enabling faster diagnosis and treatment and improving patients lives. References 1. Ong-Abdullah, Meilina, et...Biosensors Read Our Tips for Using FRET in Your Experiments Post Read Other Blog Posts about Fluorescent ...

  14. Enabling high school research at the Journal of Emerging Investigators

    Type
    Blog Post
    Published
    Sept. 5, 2018, 12:01 p.m.
    ... to teach students about basic plasmid biology Use or CRISPR 101 eBook to introduce students to this amazing...an open-access journal that enables high school students to publish peer-reviewed scientific research. ... and hear from a fantastic team of high school students who recently submitted to the journal with funding...Investigators. Chapter 2 - Getting high school students and teachers interested in JEI (the importance...audience and their current readership. Encouraging students to do science anywhere.  Chapter 4 - JEI's mission...

  15. PRIDICT: Predicting Efficiencies of Prime Editing Guide RNAs

    Type
    Blog Post
    Published
    June 25, 2024, 1:15 p.m.
    ...technology that allows precise modifications of DNA (replacements, small insertions, and deletions) without introducing...based on patterns observed from high-throughput experiments, where we can evaluate tens of thousands of designs...Figure 1). Through this method, we found key determinants of prime editing efficiency such as the length...   How to apply PRIDICT in prime editing experiments Prediction models only become useful once they...PRIDICT2.0) can greatly facilitate prime editing experiments, thus saving you time, sweat, and money on extensive...especially shines once large-scale prime editing experiments come into play. Nicolas Mathis received his ...efficiencies using sequence features and DNA repair determinants. Nature Biotechnology 2023, 1–11. https://doi.org...

  16. Viral Vectors 101: Voltage Indicators

    Type
    Blog Post
    Published
    Jan. 30, 2024, 2:15 p.m.
    ...Thankfully, there is a tool to measure these electrical events! Voltage indicators are voltmeters for cells! In...chemigenetic sensors. These tools have two main elements: a voltage-sensing component and a fluorescent...voltage sensor with a genetic component. The VSD components have historically come from a variety of sources...squirts. This design structurally separates the two components of the indicator, which can make the kinetics...types of sensors can have a variety of different components, but all contain a FRET pair, a genetic component...Viral vectors are mechanisms by which genetic components can be delivered to cells both transiently and...covered. Happy voltage sensing!   Figure 3: Components of the types of voltage indicators reviewed. ...

  17. AAV Vector Quality Control: Going the Extra Mile with NGS

    Type
    Blog Post
    Published
    Sept. 12, 2017, 1:44 p.m.
    ...are used to repeating experiments to confirm their findings. But no scientist wants to repeat an experiment...errors increases. In addition to human error, fragments of DNA from the helper plasmids or the cell genome...serotype of the packaged DNA and look for potential contaminants. The analysis is a 2-step process using Geneious...software looks for and quantifies the presence of recombinants in recombinase-dependent sequences. Recombinase-dependent...powerful tool that can be used to identify DNA contaminants in AAV preparations and provide information ... AAV vectors for your research. Please use the comments section below to let us know if you have any specific...

  18. Negative Can Be Positive: Open AAV Data with Addgene

    Type
    Blog Post
    Published
    Sept. 6, 2022, 1:15 p.m.
    ...vectors in the brain and peripheral nervous system presents a special challenge to scientists. What works ...neurons (Figure 1). However, this one set of experiments didn’t provide any information on other important... of the virus to adjacent areas impacted the experiments we wanted to perform. It required a great deal...community concerns AAV vectors are complex biological reagents; many factors, like storage buffers and conditions...conducted to improve our Addgene Data Hub, 76% of respondents indicated they wanted to see more negative data...data they cannot explain, perhaps out of fear it points to flaws or gaps in their technical ability. But...and use this data to guide them in their own experiments. By providing DOIs, this information can be cited...

  19. Lentiviral Vector Uses and Overview

    Type
    Blog Post
    Published
    May 19, 2016, 2:30 p.m.
    ...lentiviral vectors! Lentiviral vector history and components In the early 1990s, researchers developed viral...and Pol, increasing the number of recombination events necessary to create a replication-competent virus...trial with a lentiviral vector. CD4+ T-cells from patients with HIV-1 were transduced with a lentiviral vector...sequence against the HIV-1 envelope. As of 2013, 65 patients had been treated with this therapy, which lowered...beta-thalassemia. Lentiviral vectors are promising agents for cancer immunotherapy, but this research is ... vectors contain a deletion in the 3’ LTR that prevents aberrant activation of nearby genes. These safer...experiment. Let us know how you use lentivirus in the comments section below! References 1. In vivo gene delivery...

  20. Streaking for Single Colonies: The Streak Plate Challenge

    Type
    Blog Post
    Published
    March 2, 2021, 2:15 p.m.
    ... coli strains producing different pigments, and tasked the students to separate out colonies from each...second quadrant. The process continues until all quadrants are used, hopefully revealing single, isolated...incubation. The challenge is usually successful for the students. “They always get a few isolated colonies. I should...chromoprotein-expressing strains are easily amenable to other experiments to teach other aspects of molecular biology. ... do with the plasmids from Addgene is have the students transform them into different strains of E. coli...
Showing: 261 - 280 of 849 results