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We narrowed to 889 results for: ANT;

Showing: 341 - 360 of 889 results

  1. Teaching an Old DOG New Tricks: Controlling Protein Activity with GFP

    Type
    Blog Post
    Published
    Nov. 24, 2015, 3:30 p.m.
    ...for oxidizing environments to photoconvertible variants, it seems like GFP is always learning new things...frustration many researchers have experienced: wanting to ask and answer experimental questions, but not...needed to do so. Cepko lab researcher Jonathan Tang wanted to express a variety of proteins in single cell...nanobodies, the project really took off. Unlike most antibodies, nanobodies, which occur naturally in camels,...optogenetics in specific cell populations even easier. Advantages and other possibilities It’s clear that Cre-DOG...GFP-labeled lines, and they each have their own advantages. T-DOGGs can be made drug-inducible using the...proteins with a modular structure, constructing split variants should be relatively straightforward, but it is...

  2. CRISPR Meets Synthetic Biology: A Conversation with MIT’s Christopher Voigt

    Type
    Blog Post
    Published
    April 22, 2015, 2:06 p.m.
    ... controlled by regulatory networks in the plant. If we want to access that type of material - or do what... therapeutics, if you want to program a cell to identify and target a malignant cell, then you’d need ...talk about off-target interactions. That’s relevant if you want to make sure you don’t target anywhere else... parts are required to get the function that you want. The last few years, we’ve really been making tons... sophisticated. There are also proteins, like antibodies. But we are really just using what biology has...

  3. Live and Let Dye: Self-Labeling Protein Tags

    Type
    Blog Post
    Published
    April 16, 2024, 1:15 p.m.
    ...fusion protein that switches to any color or tag you want? It sounds like magic, but it’s real! In this post...photophysical properties of chemical dyes provide many advantages over fluorescent proteins, opening up new applications...HaloTag is actually HaloTag7, and recent specialized variants run up to HaloTag11. HaloTag is now the most popular...for every new-and-improved fluorescent protein you want to try, just swap in a new fluorescent ligand and...decorate synthetic viral particles with BG-linked antibodies to drive cell type-specific delivery of a viral...any cloning at all! As mentioned above, several variants of these tags have been developed over the years...are typically 230–250 aa. Smaller size isn’t a guarantee that a tag won’t disrupt the function of the target...

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

    Type
    Blog Post
    Published
    March 7, 2017, 3:30 p.m.
    ...set out to make my first CRISPR/Cas9 gene edit. I wanted to inactivate the gene BRAF (a kinase implicated...expression in A549 cells transduced with lentiCas9-blast. Anti-Cas9 Western blotting was performed on whole-cell...at the indicated multiplicity of infection (MOI). Anti-vinculin Western blotting is a loading control. ... additional 3 weeks under blasticidin selection. Anti-Cas9 Western blotting was performed on whole-cell...having active Cas9 (see tip 2). Pro-Tips 1. To quantify Cas9 activity using this assay, you really need...of the gene with multiple gRNAs, but the most important is that you could avoid potential off target artifacts...be easier to visualize and identify). If you need/want to put the digestion site in the middle of the PCR...

  5. A New Generation of Adenine Base Editors Improves Editing in Primary Human Cells

    Type
    Blog Post
    Published
    May 7, 2020, 1:15 p.m.
    ... produce 3 concurrent base edits to withstand antibiotic selection. As a result, 8 mutations in TadA could...modified ABE7.10 accordingly which resulted in variants named “ABE8.x”: “8” stands for the eighth round...x-d.  This resulted altogether in 40 new ABE8 variants. When comparing on-target DNA editing efficiency...replaced S. pyogenes Cas9 in the ABE complex with PAM-variant Cas9 proteins: the S. pyogenes NG-Cas9 (PAM: NG...PAM: NNGRRT) to create Sa-ABE8.x-m/d. With these variants, ABE8 could target many additional locations in...cells using 8 selected ABE8s and ABE7.10. They quantified genomic editing by measuring the reduction in... guide-independent off-target deamination, an important attribute for therapeutic application. ABE8s demonstrate...

  6. Plasmids 101: The Wide World of Natural Plasmids

    Type
    Blog Post
    Published
    Oct. 8, 2024, 1:15 p.m.
    ... While antibiotic resistance-mediated selection is indispensable for cloning work, antibiotics and plasmid-encoded...the Addgene Blog Plasmids 101: Antibiotic Resistance Genes Antibiotic Resistance: An Old Solution but...now used routinely in biotechnology to transform plant cells. Degradative plasmids: microbial hardiness...environments — like by living where no one else wants to. Some organisms manage this with plasmids that...created as whole-genome sequencing becomes more relevant (Garcillán-Barcia et al., 2023). In this post,... antibiotic...

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

    Type
    Blog Post
    Published
    Aug. 1, 2023, 1:15 p.m.
    ... can return multiple answers. You might want to take advantage of their save and recall functions when... with significant lot-to-lot variation. Additionally, this assembly, which used only antibiotic screening...Langhorst, B. W., Bilotti, K., Pryor, J. M., Cantor, E. J., Canton, B., Knight, T. F., Thomas C. Evans, J.,...you get results you like. NEBridge SplitSet If you want to design an overhang set specific to your DNA sequence...J. M., Potapov, V., Kucera, R. B., Bilotti, K., Cantor, E. J., & Lohman, G. J. S. (2020). Enabling one-pot...

  8. Story of a SynBio Startup: RevBio's Epiphany (or Lack Thereof)

    Type
    Blog Post
    Published
    June 5, 2014, 3:32 p.m.
    ...companies that produce GMO food plants. When a press piece ran on the detector plants, we expected to see that... lab focused on product development: building a plant that could sense and respond to novel substances...Facebook comment.  In our day job, we worked on plants engineered to detect explosives and rapidly indicate... a beautiful GMO, one that people would actually want to have. Once we had the idea, we knew it was a ...It has also allowed us to really analyze what we want out of our careers: to build something new and beautiful...we are building this company, and that’s what we want it to do. It’s exhilarating to have that sort of...have great partners in this accelerator program who want us to succeed. We’re excited to see what the next...

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

    Type
    Blog Post
    Published
    July 13, 2017, 2:30 p.m.
    ...third post will describe how you can make your own mutant flies. Find Drosophila Resources at Addgene The...combs. To make genetic crosses properly, it is important to isolate females from males while they are still... after hatching into their adult phase. It is important to clear out the parent crosses from the container...method for maintaining stable lines of genetic mutants, including in situations where maintaining heterozygosity...feature of balancer chromosomes is that they produce dominant phenotypes visible on the flies. For example, ...feature when setting up genetic crosses. More importantly, this knowledge will stop you from embarking ... of Fly Pushing by Ralph J. Greenspan. It’s a fantastic book for newbies on how to work with flies. ...

  10. Truncated gRNAs for Regulating Gene Expression

    Type
    Blog Post
    Published
    Jan. 10, 2017, 3:37 p.m.
    ... and this nuclease-null or “dead” Cas9 (dCas9) variant can then be fused to effector domains such as transcriptional...limitations to a system that relies on two distinct Cas9 variants—nuclease-competent or nuclease-null—in order to...with both the nuclease-positive and nuclease-null variants of Cas9 along with the necessary gRNAs, but it...control which gRNA becomes complexed with which variant of Cas9. To circumvent this problem, previous studies...suffers from several limitations. Perhaps most importantly, most Cas9 orthologs are less well characterized...target site. Benefits of truncated gRNAs Importantly, the use of truncated guides has largely been...graduate student at the University of California, Santa Cruz, where her research is focused on engineering...

  11. Plasmids 101: Optimizing Plasmid Yields

    Type
    Blog Post
    Published
    May 26, 2016, 2:30 p.m.
    ...strains, a change in media may be advantageous. Reduce the antibiotic concentration Low copy plasmids produce...additional steps are needed to optimize yield. You might want to include an extra wash step or use a different...suggest retransforming the plasmid into a strain meant for DNA purification. It sounds like extra work,...many high copy plasmids, standard LB broth and antibiotic concentrations will work just fine; however, ... produce fewer transcripts of their antibiotic resistance genes and can therefore be cultured in media...media containing half the normally recommended antibiotic concentration. Add supplements to the media You... Resources on Addgene.org Find Info on How to Quantify Your DNA Learn to Do a Bacterial Transformation...

  12. Tips for Using BLAST to Verify Plasmids

    Type
    Blog Post
    Published
    May 29, 2014, 1:29 p.m.
    ...differently and you may want to select one (or both) depending on the information you want to verify. We will...Organism box. This small piece of information can significantly reduce your wait time for blastn, blastp, and...expected ORFs, mutations or truncations. A primary advantage of blastx is that you do not have to decide on...using blastx at Addgene, we use the default “Non-redundant protein sequences (nr)” database as it contains...recommend using blastp over blastx. The primary advantage is time savings but an added benefit is that blastp...the default scoring matrix BLOSUM62, but you may want to check the description of the other matrices to... to see if another would be more advantageous for your search. Timesaving Tip #3: Note that protein databases...

  13. With an Eye Towards the Future, We Look Back at the March for Science

    Type
    Blog Post
    Published
    Oct. 24, 2017, 1:54 p.m.
    ...for science, and numerous other reasons. It is important to remember that we, scientists and non-scientists...role of science in our democracies will require constant vigilance and actions beyond marches, as well ...and calling for billions of dollars in cuts to important research funding.  So when I saw that there was...depends on it. Deanna McLafferty is an editorial assistant and activist. Washington, DC, USA 38.9072°N 77.0369...scientist.  I care about my work because it's important to our health.  Most people won't argue with that...been so well-publicized, we might encounter hostile anti-science activists. However, I must say that I was...I’ve worked tirelessly to get the publications, grant funding, and teaching experience that it takes to...

  14. Editor's Choice, October 2016

    Type
    Blog Post
    Published
    Nov. 4, 2016, 2:30 p.m.
    ...CRISPR in plants, to learn techniques for detecting genome edits, to discover why you might want to use ... sent Outreach Scientists to Colorado, Berlin, Atlanta, Ottawa, Norwich, and Hannover, we raised money...popular blog post this month shares incredibly important information for anyone considering a postdoc. ... through your PhDs. Of course there were many fantastic posts in October. Click on the links below to ...Advice Part 2: Building Community Engineering the Plant Genome Using CRISPR/Cas9 Healthcare Consulting: ...Door to the Business of Life Sciences 6 Tips for Grant Writing 5 Reasons to Use Reddit for Science Communication...

  15. CRISPR 101: Ribonucleoprotein (RNP) Delivery

    Type
    Blog Post
    Published
    Sept. 6, 2018, 12:02 p.m.
    ...genomic deletions. What are the advantages of Cas9 RNPs? There are many advantages to using RNPs for CRISPR ... Delivering Cas9-gRNA ribonucleoproteins  An advantage of performing CRISPR experiments using RNPs is... can be used to deliver a Cas9-gRNA RNP. This advantage has allowed scientists to perform in vitro and...et al. 2018). This was accomplished using a Cas9 mutant (Cas9M1C/C80S) that harbors two surface exposed...integration in crops, delivery of Cas9-gRNA RNPs to plants, as an alternative to plasmid based techniques,... account with these types of experiments. One important consideration is that the Cas9-gRNA RNP will be...it is likely that the use of RNPs will play an important role in its continued advancement. Andrew Hempstead...

  16. Don’t FRET: Bimolecular Fluorescence Complementation Makes Visualizing Protein-Protein Interactions Easy

    Type
    Blog Post
    Published
    Aug. 27, 2020, 1:15 p.m.
    ...in close proximity and present in large enough quantities (and the correct stoichiometric ratios) to generate...generate useable data. What’s more, measuring and quantifying FRET is often easier said than done. Fortunately...overexpression of fusion proteins it requires can lead to aberrant subcellular localization of the studied PIPs. ...protein construction, cellular expression, and quantification (Kerppola, 2006, 2013). These are described... supraphysiological expression levels. BiFC quantification This is where BiFC outshines FRET — BiFC experiments...BiFC experiments. The ideal control is a mutated variant of one or both of your PIPs in which the protein-interaction...requires that you empirically test a number of mutants to identify those that inhibit BiFC in your cellular...

  17. AAVs CREATed for Gene Delivery to the CNS and PNS

    Type
    Blog Post
    Published
    Sept. 28, 2017, 2:01 p.m.
    ...gut, and other organs to the CNS, are also an important target for gene delivery, especially for the study...Targeted Evolution. In this technique, a pool of mutant AAV capsid (cap) expressing plasmids is generated...infection and screening by PCR enrich for capsid variants with increased ability to infect the cell type...AAV.PHP.eB and AAV.PHP.S to their parental vectors. To quantify this comparison, capsids were packed with a GFP...out tables 1 and 2 below for a summary of the quantified transduction efficiency of AAV-PHP.eB and AAV-PHP.S... BP, et al. Cre-dependent selection yields AAV variants for widespread gene transfer to the adult brain...3293995. Additional Resources on the Addgene Blog Important considerations when using AAVs Learn more about...

  18. Isolating Genomic Regions of Interest with the CRISPR System

    Type
    Blog Post
    Published
    May 30, 2019, 12:44 p.m.
    ... as well as affinity purification with an anti-Cas9 antibody. These different tag systems and affinity...Using similar affinity purification with an anti-Cas9 antibody, the Li lab found a microRNA (miR483) that...transcription factor IRF-1. Immunoprecipitation, using an antibody targeting the 3xFLAG tag, allowed these researchers...this form, the CRISPR complex, consisting of a recombinant dCas9 protein and synthetic gRNA, is incubated...this technique, researchers have begun to make important discoveries in various biological fields. In 2015...chromatin by in vitro enCh IP technology using recombinant CRISPR ribonucleoproteins." Genes to Cells 21.4...

  19. Better Dyeing Through Chemistry & Small Molecule Fluorophores

    Type
    Blog Post
    Published
    Sept. 8, 2016, 2:30 p.m.
    ...from a sample. Chemical fluorophores can be substantially brighter and more photostable than fluorescent...reverting” to small molecule dyes seems daunting—no one wants to spend their days microinjecting fluorescent conjugates...systems consist of a genetically encoded enzyme variant tag that reacts specifically and irreversibly with... lipoic acid ligase, biotin ligase, phosphopantetheinyltransferase) – These enzymes catalyze the covalent...Fluorogen activating proteins (FAPs) – These modified antibody fragments bind and enhance small molecule fluorogens...excess dye from the sample, which is particularly important for samples where washing is difficult, such as...Nevertheless, these fluorophores were designed as antibody and oligonucleotide labels for fixed-cell imaging...

  20. CRISPR 101: Cytosine Transversion Editors

    Type
    Blog Post
    Published
    March 25, 2025, 1:15 p.m.
    ...between the purine bases (adenine, guanine, and hypoxanthine) or between the pyrimidine bases (cytosine, ... purine to a pyrimidine or vice-versa. Note: hypoxanthine (shown) is the nucleobase component of the nucleoside...have scientists solved this problem? By taking advantage of repair pathways within the cell — in some cases...Created with BioRender.com. Base editors take advantage of this single-stranded DNA (ssDNA) region, using...they expanded CGBE target scope by creating BE variants with other Cas9s that recognize different PAMs...processes that would cause DSBs, and its inclusion significantly reduced indels generated by CGBEs. What's next... aspiring genetic editors can now compare the advantages of base editing and prime editing, an alternative...
Showing: 341 - 360 of 889 results