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We narrowed to 839 results for: LEA

Showing: 321 - 360 of 839 results

  1. Making CRISPR Plasmids Using Fragmid

    Type
    Blog Post
    Published
    Aug. 29, 2024, 1:15 p.m.
    ...vector via Golden Gate cloning, perform an exonuclease cleanup, transform, plate, and pick. In short, the...found instead ones that were… pretty close, or at least close enough, you’ve found yourself with a common...Fragmid uses Bbsl, a Type II restriction enzyme that cleaves outside of its recognition sequence. This means...in duplicate, they were able to confirm that at least one plasmid was the correct sequence for every individual...

  2. Hot Plasmids February 2024

    Type
    Blog Post
    Published
    Feb. 20, 2024, 2:45 p.m.
    ...Cell-penetrating Cas9, fused to HIV TAT, Myc and SV40 Nuclear Localization Signals, and GFP. B) Workflow to assess...variants By Mike Lacy In 2022, Atsushi Miyawaki and colleagues identified a highly bright and stable fluorescent...just a few days later, Mohan Balasubramanian and colleagues showed the single mutation E138D is sufficient...is worth mentioning that Kiryl Piatkevich and colleagues generated yet another monomeric StayGold variant...-3188559/v1.  Neurogenetic alchemy: CAP-Mac unleashes the secrets of primate brains with brilliance ...

  3. Hot Plasmids - October 2020

    Type
    Blog Post
    Published
    Oct. 13, 2020, 1:15 p.m.
    ...ever like to write about a recent plasmid deposit please sign up here.  Here's what you'll find in this ...Caenorhabditis elegans on extrachromosomal arrays can lead to artifacts due to overexpression. CRISPR can insert...Ingaramo, Jerome Goudeau, Cynthia Kenyon, and colleagues at Calico Life Sciences. Like many split FPs,.... To resolve this issue, Shangqin Guo and her colleagues at Yale University fused a color-changing fluorescent...codon-optimized, engineered Streptococcus canis Cas9 nuclease that has a broad 5′-NNG-3′ PAM. Get the plasmids...

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

    Type
    Blog Post
    Published
    Sept. 26, 2019, 12:50 p.m.
    ...Gag-Pol polyprotein, cleaves the proteolytic site connecting the two proteins, thus releasing Cas9. Nanoblades...Additional resources on the Addgene blog Learn about RNP delivery Learn about CRISPR delivery methods for mammalian...CRISPR guide Find CRISPR plasmids and resources Learn about viral production ...

  5. Savvy Advocates Needed to Navigate a Scientific Enterprise in Flux

    Type
    Blog Post
    Published
    Sept. 13, 2016, 2:30 p.m.
    ...scientific enterprise and the way it is funded and to learn tangible skills necessary for affecting change. ...eligible for overtime pay. For many postdocs this will lead to a raise in salary, particularly given that this... a rough road ahead for students interested in leading the process despite the recent advance. Similarly... treatment of trainee scientists as workers, at least for the purposes of labor laws, rather than as trainees...Addgene's Take Aways from the FoR 2015 Symposium Learn to Advocate with the Perfect Science Talk Find a...

  6. Plasmids 101: FLEx Vectors

    Type
    Blog Post
    Published
    April 28, 2016, 2:30 p.m.
    ...previous post from our Plasmids 101 series, we learned how the Cre-loxP recombination system can be used...DNA fragment using either loxP or lox511 sites, leaving two identical sites on one end of the DNA fragment...reporter will be greatly reduced, allowing for clearer characterization of the location, morphology, and...containing FLEx technology. Read the genOway press release for more information. References: 1. Schnütgen ...Means of Genome Engineering Using FLEx AAV Vectors? Learn about Viral Vector Elements Read Our Primer on Optogenetics...

  7. Chromoproteins: Colorful Proteins For Molecular Biology Experiments

    Type
    Blog Post
    Published
    Feb. 4, 2020, 2:15 p.m.
    ...Chromoproteins also enhance corals’ resistance to mass bleaching, the loss of their symbiotic algae, during heat...rhamnose-inducible promoter. The resolution step leads to the desired mutation and excision of the vector...mammalian cell expression here   Now that you’ve learned what chromoproteins are and how they can be used...blog Find our Fluorescent Protein 101 blog series Learn more about FRET Find protocols and tips for making...Blanquet RS, Phelan MA (1987) An unusual blue mesogleal protein from the mangrove jellyfish Cassiopea ...

  8. Academic vs. Industry Postdocs

    Type
    Blog Post
    Published
    Nov. 13, 2018, 1:22 p.m.
    ...experience, I was still incredibly nervous about leaving academia. A lot of academics tend to view industry...postdoc experience, I’m going to share what I’ve learned and hopefully make your decision a little easier...publications, which may take a longer time to acquire. Learning about drug discovery: Unless your academic lab...market usually takes over a billion dollars and at least a decade of work. The number of steps, decisions...an academic postdoc, you’re often required to at least attempt to write grants. If you don’t enjoy or excel...

  9. Quick Guide to Working with Drosophila Part 1: Getting Started with Flies

    Type
    Blog Post
    Published
    July 13, 2017, 2:30 p.m.
    ...hatching into their adult phase. It is important to clear out the parent crosses from the container before...sharing-oriented, and there are many resources to help you learn more about fly biology and fly-based experimental... and gene expression. In the next post, you’ll learn the fundamentals about how many of these resources...between pathogens and their hosts. It’s not always clear which side he roots for. You can try to convince...Methods for Site Specific Recombination in Drosophila Learn about Cre-Lox Resources on Addgene.org Find Drosophila...

  10. Genome-wide Screening Using CRISPR

    Type
    Blog Post
    Published
    Aug. 18, 2015, 2:30 p.m.
    ...Identifying which genes are important for a phenotype can lead to a wealth of additional experiments investigating...consists of two components: the non-specific endonuclease (Cas9) and a single stranded guide RNA (gRNA...to enrichment or depletion of specific gRNAs can lead to skewed results. Selecting a cell type - Theoretically...validation practices, CRISPR libraries can help you learn a lot about your phenotype of interest. For more...Lentiviral CRISPR Libraries” (McDade et al., 2016). To learn a bit about how CRISPR/Cas9 can be used in other...

  11. Uncovering Ancestral Bias in CRISPR Research

    Type
    Blog Post
    Published
    Nov. 29, 2022, 2:15 p.m.
    ...dependencies  Take, for instance, a recent preprint released by the Boehm and Beroukhim labs at the Broad Institute...genetic dependencies of individual tumors, which can lead to new cancer targets and related biomarkers . Data...strongly correlated genetic dependencies that were clearly associated with European or East Asian ancestry... accounted for in the CRISPR guides they used, leading to mismatches between the guides and the lines’...undercounted by approximately 20%. In some cases, this may lead researchers to deprioritize genes that could be ...

  12. CRISPR 101: RNA Editing with Cas13

    Type
    Blog Post
    Published
    July 31, 2020, 12:30 p.m.
    ...our CRISPR Featured Topic Page Learn about RNA Targeting with Cas13a Learn about CRISPR DNA Base Editing...contain the RuvC and HNH domains responsible for DNA cleavage, so they cannot directly edit the genome. A Cas13...They hoped to find a stably folded ortholog that cleaved RNA robustly, unlike LwaCas13a, which must be stabilized... multiplexed manner to edit multiple RNAs. It’s clear that CRISPR RNA editors will open new doors for ...

  13. DNA Purification Without a Kit

    Type
    Blog Post
    Published
    Feb. 11, 2020, 2:15 p.m.
    ...cells are lysed, RNAs and proteins are removed to leave you with DNA. This DNA is then washed, and either...purify DNA, cellulose also efficiently binds, or at least entraps nucleic acids. The Botella lab took advantage...around. Finally, purified total nucleic acids are released by dipping the cellulose directly into a PCR buffer...columns, suggesting that columns can be reused at least five times. DNA purification kits don’t come with...used plasmid purification columns could also be cleaned with 1M phosphoric acid and re-used, but trace ...

  14. Introducing Addgene's New Search - Find Plasmids More Easily For Your Research!

    Type
    Blog Post
    Published
    Dec. 3, 2019, 11:38 a.m.
    ...and mascot Blugene (check back on December 5 to learn more about this!). I’ve assembled a series of screenshots...filter, this one from the ‘Requests’ category, will leave us with a single plasmid to consider. If you get...search. Simply click the ‘X’ next to any filter to clear it. Here’s a list of available filters to refine...great improvement for our website and we hope it leads you to exactly what you need with every use. If ...redesign. Additional resources on the Addgene blog Learn more about other features on Addgene's website ...

  15. CRISPR 101: Ribonucleoprotein (RNP) Delivery

    Type
    Blog Post
    Published
    Sept. 6, 2018, 12:02 p.m.
    ...your cells of interest. Cas9 RNPs are capable of cleaving genomic targets with similar efficiency as compared...levels shortly after transfection, and are quickly cleared from the cell via protein degradation pathways....known as nucleofection, which forms pores in the nuclear membrane, allowing for entry of a DNA template..... "Receptor-Mediated Delivery of CRISPR-Cas9 Endonuclease for Cell-Type-Specific Gene Editing." Journal...the Addgene Blog Subscribe to CRISPR blog posts Learn about other CRISPR delivery techniques Find tips...

  16. Better Dyeing Through Chemistry & Small Molecule Fluorophores

    Type
    Blog Post
    Published
    Sept. 8, 2016, 2:30 p.m.
    ...contributed by guest blogger, Luke Lavis, a Group Leader at the Janelia Research Campus, Howard Hughes Medical... photoactivation, etc. These impressive advances lead to an obvious question: In this age of GFP and its...fluorophore can emit a limited number of photons before bleaching. The amount of information one can extract from...Luke D. Lavis. Luke D. Lavis is currently a Group Leader at the Janelia Research Campus, Howard Hughes Medical...PMC3246539. Additional Resources on the Addgene Blog Learn About Other Protein Tags Catch Up on Your Fluorescent...

  17. Plasmids 101: Colony PCR

    Type
    Blog Post
    Published
    May 12, 2016, 2:30 p.m.
    ...Resources on the Addgene Blog Learn to do Restriction Cloning Try SLIC cloning Learn about Gibson Assembly ... One key difference is the plasmid DNA must be released from the bacteria in order to serve as PCR template...reaction. You just need to lyse the bacteria to release the plasmid DNA by either briefly boiling the sample...Read about Choosing a Molecular Cloning Technique Learn to Perform a Bacterial Transformation Deposit Your...

  18. Fluorescent Proteins 101: Green Fluorescent Protein (GFP)

    Type
    Blog Post
    Published
    May 15, 2014, 3:33 p.m.
    ...visualizing promoter activity, and much more. Read on to learn more about GFP, how scientists have evolved this...exist with only slight variations separating them.  Please note that many fluorescent proteins found on the...F64L; S65T, Y66W; N146I; M153T; V163A; A206K Cerulean F64L, S65T, Y66W, S72A, Y145A, H148D, N149I, M153T... Y145A and H148D Stabilizes the structure of Cerulean derivatives F99S, M153T, V163A Improves folding...visualize the effects of alterations to this activity. Learn more about optogenetics at OpenOptogenetics and ...

  19. 27 Hot Plasmids from 2016

    Type
    Blog Post
    Published
    Dec. 22, 2016, 3:03 p.m.
    ...different chromatin states. Alexander Stark and colleagues at the Research Institute of Molecular Pathology...Initiative, led by Dominic Esposito, has recently released a collection of 360 Ras pathway plasmids to the...Drosophila cells, targets 13,501 fly genes with at least three independent gRNAs, each chosen strategically...vacuolar) pH environments that inactivate SEP and leave only DsRed to fluoresce (1). With variants that ...effects on gene regulation. Professor Zoldos and colleagues from the Department of the Biology, Division ...26912792 FusX-TALEN assembly system TALEs as nucleases (TALENs) are genome-editing tools that are deployed...auxotrophies or using different auxotrophic backgrounds can lead to disparate physiological effects and complicate...

  20. Hot Plasmids Spring 2024

    Type
    Blog Post
    Published
    May 23, 2024, 1:15 p.m.
    ...CROPseq-multi using selection by Puromycin, Zeocin®, or nuclear expression of BFP2.     Figure 1: CROPseq-multi...separated multiple sgRNAs with tRNAs, which are cleaved out of the RNA transcript by endogenous processing...and found the protein La (a small RNA-binding exonuclease protection factor) promotes editing (Yan, J et...hypothesized that the improved efficiency is at least in part due to La binding and stabilizing the 3’...Neurodegeneration Collection updates Last but not least, check out the new and noteworthy tools featured...

  21. Viral Production at Addgene

    Type
    Blog Post
    Published
    Oct. 24, 2023, 1:15 p.m.
    ...each viral lot. To learn which specific QC experiments were performed on your lot, please contact us. Full...medium (OptiPro SFM) containing lentivirus is first cleared by low-speed centrifugation and then by filtration...responsive element (RRE) gene and the cellular ribonuclease P/MRP 30 kDa subunit (RPP30) as a control for...the sample from the DNA tube all the way to the cleared viral preparation pool. In addition, the final ...

  22. CRISPR-based Genome Editing Systems For Editing in E. coli

    Type
    Blog Post
    Published
    April 13, 2021, 1:15 p.m.
    ...replicon of pTargetF may exhibit a more serious leaky expression in BL21(DE3), which we presume caused...that indeed the gRNA-pMB1 had a slightly higher leaky expression in BL21(DE3) that resulted in a lower...a sacB gene renders it beneficial for plasmid clearance. It is more convenient to update the spacer(20nt...Anhydrotetracycline (aTc) induces expression transposases leading to increased integration frequency. (C) Increasing...rounds of transfer on transposition-inducing agar lead to higher cargo copy numbers. (D) The effect of ...

  23. Deciphering the Mysteries of Behavior with Viral Vectors

    Type
    Blog Post
    Published
    Sept. 17, 2019, 12:40 p.m.
    ...integrating neuronal signalling and triggering release of the stress hormone norepinephrine to promote...are chemogenetically activated, norepinephrine release causes a freezing response, a form of immobility... the NAc, where enhanced glutamate transmission leads to locomotion sensitization. (for review see van... as sleep, food intake, mobility/coordination, learning, and memory. We’ll cover these in a  future blog...Guide to Viral Vectors Resources on Addgene.org Learn about Addgene's viral service Find viral protocols...

  24. Beyond the Blood Brain Barrier: PHP.B Enters the Central Nervous System Via the LY6A Receptor

    Type
    Blog Post
    Published
    July 23, 2019, 12:46 p.m.
    ...tips from an AAV rookie  Learn about some considerations when using AAVs Learn more about using AAV as ...complex (LY6A) as a receptor for PHP.B. Read on to learn how the design of PHP.B may have contributed to ...182-188. PubMed PMID: 29175632. If you want to learn more about how PHP.B and PHP.eB were engineered,...PMC5088052. Additional resources on the Addgene blog Learn more about PHP.eB, and PHP.S, a peripheral nervous...

  25. The Challenges of Cell Culture

    Type
    Blog Post
    Published
    May 5, 2016, 2:30 p.m.
    ...Blog Learn All about Mammalian Vectors Read up on CRISPR Delivery Systems for Mammalian Cells Learn about...older than those available from biorepositories leading to genotypic and phenotypic artefacts. The life...commercial repositories and the same cell line from a colleague?”. The main difference is the QC and validation...usually the case if you obtain a cell line from a colleague. Time and money are wasted, papers are retracted...

  26. Avoiding the Dark Side of Fluorescent Protein Fusions with mOX FPs

    Type
    Blog Post
    Published
    Oct. 27, 2015, 3 p.m.
    ...remains unclear whether the FP misfolding impacts fusion protein function and this could lead to under...localize [4]. Most investigators are surprised to learn that FPs are often poorly suited for cellular compartments...glycosylation, disulfide bond formation, and proteolytic cleavage. FPs localized to organelles are equally susceptible... dark fluorescent proteins in your experiments, please consider the following: How can one ensure that...

  27. Important Considerations in Optogenetics Behavioral Experiments

    Type
    Blog Post
    Published
    Oct. 1, 2015, 2:30 p.m.
    ...bit of a steep learning curve and many opportunities for mistakes. The easiest way to learn optogenetics... is an approximation of reward or the sense of pleasure that the animal perceives through taking a drug...University.  His interests include the role of learning and memory in opioid addiction and science communication...Kheirbek, Mazen A., et al. "Differential control of learning and anxiety along the dorsoventral axis of the...

  28. Welcome to the Lab Bench! New Protocol Series Helps Trainees Get Started in the Lab

    Type
    Blog Post
    Published
    Sept. 7, 2021, 1:15 p.m.
    ...Bench series was born from the shift to remote learning due to COVID-19. We had many protocols for plasmid...details about personal protective equipment to learning about how to use a tissue culture hood and other...other lab equipment, we’re here for you as you learn the ropes! All material can be found on our Protocols...Protocols page and YouTube page.  Happy learning, and welcome to the lab bench, new scientists!   ...

  29. Plasmids 101: Expressing shRNAs for RNA Interference

    Type
    Blog Post
    Published
    June 17, 2025, 1:15 p.m.
    ...which is then cleaved by RISC and degraded. In the miRNA pathway, RISC doesn’t always cleave the mRNA — ...hairpin shRNA structures that are processed by a ribonuclease called Dicer. In a great simplification of the...protein is what binds the small RNA and either cleaves the target mRNA or recruits other silencing proteins...no tissue specificity, so in some cases they can lead to toxicity and off target effects. More recently...

  30. Viral Vectors 101: Integrase-Deficient Lentiviral Vectors

    Type
    Blog Post
    Published
    May 13, 2025, 1:15 p.m.
    ...investigating HIV integrase mutants in the early 1990s (Leavitt et al., 1996), and since then, numerous variants...However, IDLV genomes are transcriptionally active, leading to transient expression of gene products (Wu et... insertional mutagenesis. This integration could lead to an undesirable interruption of a critical gene... off-target mutagenesis induced by CRISPR-Cas nucleases in human cells. Nature Biotechnology, 31(9), 822...PMID: 18688286 PubMed Central PMCID: PMC2496964 Leavitt, A. D., Robles, G., Alesandro, N., & Varmus, H....

  31. The Fluorescent Vegetables in Aptamer Soup

    Type
    Blog Post
    Published
    Jan. 29, 2019, 2:27 p.m.
    ...warm meal on a winter day, you should be happy to learn that many DNA-based reagents are named for soup ... and non toxic to cells. Because it rapidly photobleaches, DFHBI can only be used for qualitative, but...regions in the 5’ UTR of mRNA that bind metabolites, leading to conformational changes that control the expression...the Corn-DFHO complex. For dessert (last but not least), the only fruit-named aptamer, Mango, binds to ...PMC4143336. Additional resources on the Addgene blog Learn more about aptamer fluorophores Investigate other...

  32. Plasmids 101: Biotinylation

    Type
    Blog Post
    Published
    Nov. 15, 2018, 1:50 p.m.
    ...the uses of biotinylation in molecular biology! Learn about in vivo biotinylation of bacterial fusion ...biotin-avidin complexes bound to your protein of interest leading to amplification of the reporter signal and easier...microenvironments. There are also reversible and cleavable biotinylation reagents to help with the specific...vivo biotinylation of bacterial fusion proteins Learn how biotin is used in Split-BioID: An improved method...method for studying protein-protein interactions Learn how biotin can be used with CRISPR Additional resources...

  33. Plasmids 101: Common Lab E. coli Strains

    Type
    Blog Post
    Published
    Nov. 7, 2014, 2:56 p.m.
    ...expressed proteins endA, endA1 Endonuclease I (nonspecific cleavage of dsDNA ) mutation Improves plasmid...cell mcrA, mcrBC Inactivation of pathway that cleaves methylated cytosine DNA Allows for uptake of foreign... mrr, Δ(mcrC-mrr) Inactivation of pathway that cleaves methylated adenine or cytosine DNA Allows for uptake...recombination, increases plasmid stability recBCD Exonuclease V activity abolished Increased plasmid stability...

  34. We’re Going Viral This Week!

    Type
    Blog Post
    Published
    Oct. 14, 2019, 1 p.m.
    ...blog posts Learn more about AAVs Resources on the Addgene website Visit the AAV Data Hub Learn more about... serotype, injection site, and incubation time. Learn more about the Data Hub in this blog post. Alongside...attending SfN2019, stop by booth #2163 to say hi and learn more about Addgene’s viral service. Additional ...

  35. Recombinase-based State Machines Enable Order-dependent Logic in vivo

    Type
    Blog Post
    Published
    July 28, 2016, 2:30 p.m.
    ... the Addgene Blog Learn How Piggy Bac Vectors Can Be Used for Genome Editing Learn about Michael Koeris...needs. For more context, exposition, and detail, please refer to the paper. Why might one be interested...uploaded to Addgene. We also recommend inserting self-cleaving hammerhead ribozymes directly upstream of the ... transcribed gene and its 5’ UTR. Various self-cleaving hammerhead ribozyme parts are described in Nielsen...

  36. Addgene’s New Way to Help You Share Science

    Type
    Blog Post
    Published
    Nov. 14, 2013, 4:10 p.m.
    ... I had found my dream job, because one thing I learned in grad school was that scientists have amazing...at their lab benches. It’s these exchanges with leading scientists (and our growing collection of 25,000...sciences research for the past decade. We’ve also learned along the way that a connected and collaborative...scientists who have deposited plasmids at Addgene.  Learn how discoveries were made and get practical tips...

  37. Ghanaian Researchers to Receive Plasmid Awards from Addgene and Seeding Labs

    Type
    Blog Post
    Published
    June 15, 2016, 1:30 p.m.
    ...Health and Allied Sciences in Ghana. Read on to learn more about doctors Bengyella and Duedu and how they...tumors is “a major public health problem, and a leading cause of hospitalization for gynaecologic disorders...Program Additional Resources on the Addgene Blog Learn about Plasmid Awards to Researchers Working on Rare... in China Additional Resources on Addgene.org Learn About Addgene's Mission Check Out Our International...

  38. Important Considerations When Using AAVs

    Type
    Blog Post
    Published
    June 13, 2017, 2:30 p.m.
    ...alcohol-based disinfectants. A 10% bleach solution should be used to clean AAVs from workspaces. Speed...Manitoba due to their Biosafety Level-2 or 2+. Please Note: Despite the fact that AAVs themselves are...vectors (e.g. Lentivirus, Adenovirus) AAVs cause the least amount of immune response. Any investigator who ...antisense oligonucleotides (ASOs), zinc finger nucleases, and many others (12). The most attractive potential...

  39. Quick Guide to All Things Lentivirus

    Type
    Blog Post
    Published
    March 21, 2017, 2:30 p.m.
    ...LTRs) has been removed, Rev (which facilitates nuclear export) is expressed from a separate plasmid, and...mutation or recombination with human retroviruses could lead to RCL. Regarding transduction efficacy, no differences...Quickly after transfection (4-8h), cells start releasing viral particles into the culture supernatant and...retrovirus packaging cell lines to avoid recombination leading to helper virus production." Molecular and Cellular...5 Tips for Trouble Shooting Viral Transductions Learn Why You Might Want to Use AAV in Your Research Lentiviral...

  40. Antibodies 101: Validation

    Type
    Blog Post
    Published
    March 24, 2022, 1:15 p.m.
    ...even to not bind to their target molecule at all, leading to inconclusive results, erroneous conclusions,... that your antibody has been validated using at least one of these strategies. Choosing which strategy...) the protein of interest in your sample, should lead to reduced or eliminated antibody signal. If you...translation start sites or alternative splicing that could lead to some form of the target protein to be present... Check out our Selecting an Antibody blog post! Learn more about Antibodies and their many applications...
Showing: 321 - 360 of 839 results