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We narrowed to 1,028 results for: ELL

Showing: 521 - 560 of 1028 results

  1. Quick Guide to Working with Drosophila Part 2: Controlling Gene Expression in Flies with Gal4/UAS

    Type
    Blog Post
    ... to express one gene in one cell type and another gene in a different cell type (6–8). Although it’s often...systematically study gene function with temporal control and cell-type specificity! The Gal4/UAS system At a more...the hedgehog gene can be overexpressed in several cell types by using different Gal4 drivers. It is much...much easier to mate flies to get the right cell type expression as opposed to making a new fly with hedgehog... Targeted gene expression as a means of altering cell fates and generating dominant phenotypes., Development...PubMed PMID: 15262411. 4. T. D. Southall, D. A. Elliott, A. H. Brand, The GAL4 System : A Versatile Toolkit...Neuro-Trauma during Asymptomatic Viral Infections, Cell Host Microbe 22, 48-60 (2017). PubMed PMID: 28704652...

  2. Fluorescent Protein Travel Awards - Protein Variants, a Serotonin Sensor, and an Artificial Leaf Replica System

    Type
    Blog Post
    ...non-growing cells are orange, slow growing cells are orange-green, and fast growing cells are green. A...fluorescence. Cells expressing the library are then sorted by fluorescence-activated cell sorting into ...for serotonin. According to Unger, “serotonin is a well-known but poorly studied neuromodulator, and evidence... A second colonizer, labelled with a constitutively fluorescent reporter can also be used to track how...population. Last, Bernach will simultaneously monitor cell cycle and growth rate using TIMER and an unstable...reporter under the control of promoters that control cell cycle gene expression.  Additional Resources on...

  3. Fluorescent Proteins 101: Luciferases

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    Blog Post
    ...Susceptible Subcellular imaging Improving Well-established High-throughput assays Available Well-established...luciferase assays require cell lysis as the most efficient means to disrupt the cell membrane and deliver ...catalyzes a chemical reaction in its abdomen to produce yellow-green light (Figure 2). Despite the title of this...produced. A similar approach can be used to analyze cell signaling pathways or miRNA knockdown. BRET for ...advantageous properties. This approach can improve live-cell imaging by avoiding autofluorescence and phototoxicity...can permit measurements of luminescence from live cells. Luciferase variants with incompatible substrates...

  4. Flow Cytometry Readouts: Yes, No, and Everything in Between

    Type
    Blog Post
    ... plot would serve you well. Percentage of cells in a parent gate (all live cells for example) that are...percentage of total live cells are GFP+. In this plot, with adjusted voltage, cells that did not receive ...example, you have introduced a GFP protein into your cells (visualized via the FITC channel) and you are trying...graphs could simply be the percentage of the total cells present in the gate.   Figure 1: Example...looking at proteins that are expressed regularly in cells and might be upregulated or downregulated based ...populations. MFI is the average brightness from all cells that are positive for the marker of interest. MFI...

  5. Viral Vectors 101: Viral Vector Elements

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    Blog Post
    ...genetic cargo, typically an expression construct, to cells/organisms. This is often cDNA, shRNA, or something... goal, a promoter should flank this component as well. On both sides of this promoter-gene combo are two...repeat regions in addition to their unique regions as well, though. Viral packaging and entry The genetic cargo...perform very similar functions to infiltrate the host cell and deposit viral cargo. Gene expression While AAV...what will be delivered and expressed in the host cell from the transfer plasmid. Viral packaging and entry...altered tropism of AAV to target different tissue or cell types. AAV also requires some back up from adenovirus...other. Below we will outline the common features as well as the differences within their viral elements. ...

  6. Antibodies 101: ChIP

    Type
    Blog Post
    ...spin down the samples to remove cell debris, which will collect in a pellet. The supernatant will contain... you may be thinking, well, these are mostly fairly standard molecular and cell biology techniques. What... Process ChIP can be done for either tissues or cells. The process is very similar for both, though tissue...Both polyclonal and monoclonal antibodies can work well in ChIP. If you’re using an antibody for the first...What’s all the fuss? Well, ChIP’s complexities come not from the techniques used, but from the difficulty...will also have to be optimized for both tissue or cell type, amount of protein, and desired fragment length...other antibody applications, this investment can well be worth it once you generate your data. However...

  7. A Novice's Perspective on Neuroscience - SfN2017

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    Blog Post
    ...aiming to dedifferentiate mouse brain cells into neural stem cells. I was quite unsuccesful (although I... work in sociology, behavior, and artificial intelligence (including a team that created a novel form ...game Go). In addition, there was an entire story-telling session organized by Story Collider and which featured...neuroscience trade. There were, of course, many companies selling antibodies, microscopes, and dissection tools, ...versatile means of delivering genes to many different cell types with minimal immunological effects. AAVs are... used in experiments targeting genes to specific cell types in the brain, mapping neuronal networks, and...Addgene booth to learn more about these tools and to tell us how useful the service has been for their experiments...

  8. Q&A with Karen Katz and Carl Paratore, the Newest Members of the Addgene Board of Directors

    Type
    Blog Post
    ... shared. Can you tell me a bit about your job? Katz: I’m the Director of Intellectual Property Programs... Board of Directors. Katz is the Director of Intellectual Property Programs at Suffolk University Law ...provides interesting and unique challenges for intellectual property lawyers. While there will always be...there will always be an interest in protecting intellectual property. Figuring out that balance is, and ...Suffolk is to help create new programmatic content as well as help our students succeed whether that be academically... Karen Katz, Esq Ms. Katz is the Director of Intellectual Property Programs at Suffolk University Law ...she was on law review. She clerked for Juan R. Torruella, U.S. First Circuit Court of Appeals. She served...

  9. 15 Years of Addgene: The Top 15 Plasmids

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    Blog Post
    ...plasmid. Envelope proteins determine which types of cells a virus can infect. The VSV-G envelope protein enables... enables this virus to infect many different cell types so it is useful in a wide range of applications... The 2A-GFP allows for screening of transfected cells. Find pSpCas9(BB)-2A-GFP (PX458). lentiCRISPR v2...deliver Cas9 and your gRNA of interest into mammalian cells. This vector was the top requested plasmid of 2015... popular in genome engineering. For use in human cell lines, the Zhang lab codon-optimized SpCas9. Using...puromycin cassette that makes it less efficient in some cell lines. pSpCas9(BB)-2A-Puro (PX459) V2.0 was made...capable of inducible expression of shRNA in target cells by the addition of tetracycline/doxycycline to the...

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

    Type
    Blog Post
    ...Ishitani R, Zhang F, Nureki O. Cell. 2014 Feb 27;156(5):935-49. doi: 10.1016/j.cell.2014.02.001. Epub 2014 Feb...Kampmann M, Weissman JS. Cell. 2014 Oct 23;159(3):647-61. doi: 10.1016/j.cell.2014.09.029. Epub 2014 Oct...Can you engineer CRISPR-Cas9 activators that work well enough on any gene that they could be used by people...screening for genes that, when turned on, make melanoma cells resistant to treatment with a BRAF inhibitor. (The...Addgene, enabling systematic investigation of the cellular consequences of repressing or inducing individual...Weissman and his colleagues demonstrated in a paper in Cell that they could typically achieve 90-99% knockdown...straightforward to apply the new library for use in various cell types and in live mice. Therapeutic uses in humans...

  11. Plasmids 101: Restriction Cloning

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    Blog Post
    ...use electro-competent cells instead of the more common chemically-competent cells. You should perform, ...This way you can then cut the plasmid backbone as well as the insert with EcoRI and HindIII and, when you...and verification required for the process to work well, so let’s go over the details step-by-step. 1. ...varies for different enzymes. Many companies now sell fast digest enzymes that can digest large amounts...backbone from the enzymes used to digest them as well as any pieces cut out or off of them. An easy way...undigested controls) are loaded at the top of the gel in wells positioned toward the cathode (- charge). When the... manufacturer’s instructions for your competent cells. For most standard cloning, you can transform 1-...

  12. RNA Extraction Without A Kit

    Type
    Blog Post
    ...are commonly found in cells, and without something to block RNase activity upon cell lysis, RNA is degraded... within cells or tissues- they don’t actually chemically stabilize the RNA molecules (Allewell and Sarma...RNases such as RNaseZAP. When harvesting tissues, cells, plants, fungi, or bacteria, keep samples cold and...before RNases had a chance to break down RNA upon cell lysis.  In the late 1970’s, Chirgwin and colleagues...keeping your samples cold until homogenization and cell lysis. This can be challenging depending on your...developed ways to “stabilize” RNA within tissues before cell lysis. These products, namely RNAlater®  from Thermo...is not an issue.  References                 Allewell NM, Sama A (1974) The effect of ammonium sulfate...

  13. RANbodies: Reporter Nanobody Fusions

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    Blog Post
    ...protein of interest in cells and tissues. Although antibody production is well established, it’s also .... RANbodies are expressed in 293T cells and are secreted from cells due to their IgGκ signal peptide sequence... sequence, making them easy to harvest from cell culture media. Their His tags allow for further purification...RANBody While all three reporters stain cultured cells and tissue sections, some reporters are better suited... signal amplification of HRP Staining cultured cells and tissue sections Amplifying weak signals of fluorescent...Labeled anti-HA or -Myc antibody Staining cultured cells and tissue sections When detected with a labeled...Labeled Anti-Chicken IgY antibody Staining cultured cells and tissue sections Great for multicolor staining...

  14. Uncertainty about Labor Law Brings More Uncertainty to Postdoc Wages

    Type
    Blog Post
    ...Many thanks to our guest blogger Gary McDowell! Gary S. McDowell is currently the Executive Director of...This post was contributed by guest blogger, Gary McDowell, executive director of Future of Research. On ...opportunity of the lapsed mandate (or are being compelled at the state level, in the case of some public...expecting a salary increase have had that increase cancelled. Rounding out the remainder, 2% of postdocs who... are continuing with their plans, and 17 have cancelled their plans. 2 of those raising - the University...Illinois and the University of Michigan - initially cancelled their plans, but have since reversed their position... In addition, PIs at some of the institutions cancelling plans are going ahead with raises anyway, including...

  15. Hot Plasmids - March 2019 - Anti-CRISPR, 2in1 Cloning, Fluorescent Voltage Indicators, and Photoswitchable Proteins

    Type
    Blog Post
    ...indicator. Archon1 can embed itself into the cell membrane where cell voltage can then be measured. Archon1 ... it. Using a robotic picking approach to select cells with variant characteristics they wanted, the team...2in1 vector cloning system Article contributed by Michelle Cronin  Listen to the 2in1 cloning podcast segment...through recombination. The 2in1 cloning system is well suited for protein-protein interaction (PPI) studies...offers a way to spatially and temporally control cellular processes in vivo. Many photoactive proteins exist...approach to control protein-protein interactions in cells because the protein binders can be customized for...photo-reversible protein-protein interactions can control subcellular localization processes in vivo. They deposited...

  16. RaPID Detection of RNA-protein Interactions

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    Blog Post
    ...designed an RNA bait containing three well conserved but not well characterized RNA motifs. This bait identified...proteins that bind an RNA motif of interest in living cells. Challenges with existing RNA-protein interaction...and naturally occurring complexes present in the cell, it can also introduce non-physiological RNA-protein...a short timescale, such as during drug action or cell signaling. Limitations of RaPID Like other BirA*... controls include both scrambled RNA motifs and cells transfected with BirA* protein alone (i.e. no RNA...protein that’s highly expressed in neural progenitor cells. Knock-down of QKI decreased Zika viral RNA levels...2018). RNA–protein interaction detection in living cells. Nature Methods, 15, 207-212. PubMed PMID: 29400715...

  17. Plasmids 101: Modular Cloning

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    Blog Post
    ...complex cellular behaviors requires making complex manipulations to cells. For example: in a cell there ... yield a cellular phenotype, translational output, etc., which can support a larger cellular engineering...TU, or even an individual part, will behave in a cell. For unpredictable cases, MoClo provides two answers...allow you to study natural genetic circuits in the cell. Natural and synthetic product production A major...biofuels, drug precursors, etc., microbially or in cell-free systems. These pathways typically involve multiple... be used to assemble multiplexed gRNA systems as well as additional components such as selectable markers...

  18. Lab Automation at Addgene

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    Blog Post
    ...protocol, we can prep two 96 well plates in just under 3 hours or one 96 well plate in about an hour. Automating...our set up. The cooling block is used to keep our cells from warming up too much and dying.  Also, we added...added a couple of platforms that fit 48 well q- trays allowing us to do up to 96 transformations in one...simple. By clicking a few buttons you can quickly tell the Biomek FxP how much sample to pick up, where...pick up 96 tips for easy sample transfer between 96 well plates. The second pod can pick up 8 tips and position...be useful if you’re transferring samples from 96 wells plates onto agar q-trays where the samples must ...Have you ever wondered who fills all those vials? Well, it used to be a lab assistant, but with increased...

  19. Science communication: 9 Strategies to Get Your Foot in the Door

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    Blog Post
    ...volunteers as well. 4) Blog - Hone your writing skills Take the DIY approach and start a blog. Stellar talents...contributed by guest blogger Sarah Schmidt, a Marie Curie Fellow at The Sainsbury Laboratory. Are you a science ... probably write for your institute’s blog. The Wellcome Trust blog or the John Innes Student Voice are...decide to start your own blog, check out these two excellent articles: On the Origin of Science Writers How...). A great opportunity is the AAAS Mass Media Fellowship, which places science students for 10 busy weeks...American or National Geographic. Similarly, media fellowships from the British Science Association allow you...started opening up. My current job as a Marie Curie Fellow is a direct consequence of a blog that I started...

  20. CRISPR Activators: A Comparison Between dCas9-VP64, SAM, SunTag, VPR, and More!

    Type
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    ...Control of Gene Expression. Cell 152:1173–1183 . https://doi.org/10.1016/j.cell.2013.02.022 Sajwan S, Mannervik...and Fluorescence Imaging. Cell 159:635–646 . https://doi.org/10.1016/j.cell.2014.09.039 Zhou H, Liu J,...from when looking to activate genes across diverse cell lines.   dCas9-VP64 Description CRISPR activation...large and are not expressed consistently throughout cells. VPR Description VPR fuses a tripartite complex ...activator used in other CRISPR activation methods, as well as two other potent transcriptional activators (...activation levels than SAM in some loci across Drosophila cell lines (Sajwan et al., 2019). SPH SPH combines components... Gabrielle Clouse...

  21. Adapting Toehold Switches to Detect Zika Virus

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    Blog Post
    ...for pathogen detection In 2014, as postdoctoral fellows with Jim Collins and Peng Yin at the Wyss Institute...circuits to be deployed out of the laboratory in a cell-free and biosafe format that was stable for up to...possible and submitted our manuscript to the journal Cell just six weeks later. The Zika publication timeline...switch designs, CRISPR/Cas9-mediated genotyping, as well as our electronic reader that provides portable ... demonstrated our technology on active virus, as well as shown that we can detect the virus in a clinically...of Toronto. His research is focused on developing cell-free synthetic biology applications for human health...Virus Using Programmable Biomolecular Components." Cell (2016). PubMed PMID: 27160350. Find Plasmids from...

  22. Reaching out to China: Canton Nucleic Acids Forum (CNAF) 2015

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    Blog Post
    ...alone. At least I could tell that they were mushrooms. In some cases I couldn’t tell if the bags contained...origin of cellular life. His lab has long hypothesized that life started with RNA and a simple cell membrane...Chinese medicine. There were hundreds of stalls selling a myriad of items, only a small number of which...by Nobel Laureate (and Addgene depositor) Craig Mello. Taking his groundbreaking work in RNAi one step...involved in preventing silencing of self-genes. Dr. Mello presented a hypothesis that these pathways are involved... circular RNA they have identified resulted in a cell migration defect. They have created a circRNA website...discover circRNAs in RNA-seq data. As far as I can tell, we don’t really know what circRNAs do yet or how...

  23. Illuminating Epigenetics with A FRET Based Biosensor

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    ...genes should be ‘read’ by the cellular machinery. This is how most cells in the body contain the same ...complex of DNA and histone proteins that compacts cellular DNA into chromosomes and reduces the risk of dangerous...histones changes, then gene expression can change as well. The more frequently and tightly bound DNA is to...the more compacted it becomes. This prevents the cellular machinery from physically interacting with the...function. Angelman syndrome is characterized by intellectual and developmental disability, jerky movements...visualise changes in histone methylation in living cells. Essentially this reporter construct consists of...recognise lysine 27 methylation. When tested in living cells where methylation of these sites was controlled,...

  24. Plasmids 101: Using Transposons in the Lab

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    Blog Post
    ...vitro as well as in vivo in yeast, plant, insect, and mammalian cells, including human cells. piggyBac...is active in vertebrates and integrates in human cells at rates similar to retroviral vectors. The hyperactive...piggyBac has an integration efficiency in human cells similar to retroviral vectors. The hyperactive PB...transposase (hyPB) is ~10-fold more active in mammalian cells compared to the codon-optimized wild-type piggyBac...G/C)TNA. Tol2 can deliver 10-11 kB to mammalian cells without decreased efficiency, with a maximum cargo...and has lower efficiency of integration in human cells than piggyBac and Sleeping Beauty. Minimal Tol2 ... or barcodes. When transposons are delivered to cells or a model organism, they insert into the host’s...

  25. Troubleshooting and Optimizing a Western Blot

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    ...check its subcellular location and solubility. If it’s membrane-bound or located in a subcellular compartment...reagent. Be aware that this approach requires excellent attention to detail, since you’ll end up with ...question before running one.  Good on gradients? Excellent! Now, let’s head west, intrepid reader! Protein...strengths. If that’s not working, consider using a subcellular fractionation kit or a lysing protocol developed...samples and reference ladders are not next to empty wells. Most people will simply load an extra ladder or...replicate (ie, leftover sample) to fill the relevant wells. Similarly, if you load a gel and then forget to...the time with smaller tasks, or simply enjoy some well-deserved down time while waiting for your timer ...

  26. Of Myc and Men

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    ...For about as long as the disciplines have existed, cell and molecular biologists have been continuously ... peer into the mysterious, microscopic world of cells. The development of antibodies as tools allowed ...was the existence of high quality antibodies for well defined peptides. Munro and Pelham’s first published...monoclonals would help the field explore Myc’s role in cell proliferation and tumorigenesis.  Using two different...the same antibody developed almost 40 years ago? Well…kinda. In some cases, antibody providers are distributing... human IgG2, mouse IgE, goat IgG, (and more!) as well as the original mouse IgG1. And the refinement of...specific for human c-myc proto-oncogene product. Mol Cell Biol 5:3610–3616. https://doi.org/10.1128/mcb.5.12.3610...

  27. Your Top Requested Plasmid in 2016!

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    Blog Post
    ...both SpCas9 and your gRNA in a variety of mammalian cell types. Purpose  SpCas9 This plasmid expresses Streptococcus...the Zhang lab has tested in a variety of mammalian cell lines. Cas9, of course, is the key component of ... puromycin resistance gene enables selection of cells with the plasmid post transfection. The 2A "self...can be used to transfect a variety of mammalian cells and can be modified for viral trasduction (see the...resistance gene that made it less effective in certain cell lines. This point mutation has been corrected in... your gRNA of interest in a variety of mammalian cell lines. Alternative forms of this vector contain ...directed repair. You can find all of these variants, as well as links to the Zhang Lab's Nature Protocols Publication...

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

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    Blog Post
    ...scientists. What works well in one cell or neuron type may not work well for a neighboring cell type, even within...options while AAV8 (82%) and AAV9 (84%) were both excellent at infecting our target neurons (Figure 1). However... conditions (3,5,8), injection titers (1,2), and cell types used for packaging viral particles (6), can...methods and image descriptions, a minimum n = 2, and well labeled high- and low-magnification images demonstrating... vector performance in various brain regions and cell types. Power in numbers A Pubmed search for “AAV....169 Gruntman, A. M., Su, L., Su, Q., Gao, G., Mueller, C., & Flotte, T. R. (2015). Stability and compatibility...

  29. The PAM Requirement and Expanding CRISPR Beyond SpCas9

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    ...Class 2 CRISPR-Cas System. Cell 163:759–771 . https://doi.org/10.1016/j.cell.2015.09.038 Find plasmids...are capable of cleaving genomic DNA in mammalian cells and zebrafish embryos, and they can be used to modify...domains and allows for expanded PAM recognition as well as increased specificity and lower off-target activity... adapted for targeted RNA cleavage in mammalian cells. The REPAIR (RNA Editing for Programmable A to I...Han S, Joung J, Belanto JJ, Verdine V, Cox DBT, Kellner MJ, Regev A, Lander ES, Voytas DF, Ting AY, Zhang... DBT, Gootenberg JS, Abudayyeh OO, Franklin B, Kellner MJ, Joung J, Zhang F (2017) RNA editing with CRISPR-Cas13...reveals specificities of Cpf1 endonucleases in human cells. Nat Biotechnol 34:863–868 . https://doi.org/10.1038...

  30. 5 Reasons to Use Reddit for Science Communication

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    ...blog The blog has many other excellent posts on science career topics as well. Instead of rewriting or coming...everyone, particularly if you’re more interested in selling a product than communicating your ideas with people...has great questions! 2. The r/Science subreddit is well moderated. One of our original worries about sharing...rousing the trolls. 3. The AMA process was simple and well organized. You can check out the r/Science AMA Submission...with our general mission to share- as opposed to sell- science. 4. You can prepare answers beforehand....answers off the cuff, but like to be able to provide well thought out responses. I hung out with Joanne as...small quibble. If you are not too worried about selling yourself and really do just want to get information...

  31. Plasmids 101: What is a plasmid?

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    ... small circular piece of DNA found in bacterial cells, and someone new to plasmids may need some extra...and designed to introduce foreign DNA into another cell. Minimally, lab-created plasmids have an origin ...the ability of plasmids to self-replicate within a cell make them attractive tools for the life scientist...component for expression vectors: determines which cell types the gene is expressed in and amount of recombinant...plasmids also have selectable markers for use in other cell types. Primer Binding Site A short single-stranded...newly inserted gene is transformed into bacterial cells and selectively grown on antibiotic plates.  Importantly...plasmids to manipulate gene expression in target cells. Characteristics such as flexibility, versatility...

  32. Management for Scientists: Giving Feedback

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    Blog Post
    ...aspect of being a successful manager. Great bosses tell people where they stand clearly and routinely. They... are clear with each employee about what they do well and where they need to improve, and they’re also...making it a concrete goal to say “thank you, job well done” to one of your employees every day. Prepare... is usually good. Don’t pretend something went well when it didn’t… but don’t “punish” someone for a...say worked out best? What 3 things worked least well? Manager provides feedback (may be the same as ...best.  Here are the 3 things that worked least well. Find out the Effect of the Feedback: How does...this feedback compare with your own perception? Tell me your interpretation of my feedback to you. What...

  33. March for Science

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    Blog Post
    ... details for the march in Washington as well as the satellite marches. Anyone can join the marches or ... by Addgene or its staff. In Washington, D.C. as well as sister locations on April 22, 2017, scientists...government protects (and often funds [3]) scientists as well as other citizens. Thus civilization proceeds in...lists over 200 partner organizations, and 514 satellite marches are scheduled to take place around the... a success. Below are resources for the march as well as other science advocacy opportunities. Information...conversation about science’s role in government as well as active advocacy will be required. Information...Website The Washington Post: Trump administration tells EPA to freeze all grants, contracts The Guardian...

  34. Harnessing Bacterial Toxins for Allelic Exchange

    Type
    Blog Post
    ... that converts sucrose into a toxic metabolite. Cells that have undergone a double crossover will not ...contain sacB, allowing them to grow on sucrose. Cells that have only gone through the single cross over...counter-selection escape). The rhamnose induction system is well-characterized and quite tight (Giacalone et al.,...., 2006). The Chen lab demonstrated excellent counter-selection which enabled very efficient recombineering...desired deletion directly into cut vector with excellent efficiency using a commercial isothermal assembly...Enterobacteriaceae including in E. coli O157:H7, Shigella flexneri, Enterobacter cloacae, and Serratia marcescens...marcescens and have created dozens of deletions as well as insertions with this system (Lazarus et al., ...

  35. Meet the New Addgene Board Members!

    Type
    Blog Post
    ...read some of the seminal papers on cell cycle discovery from Lee Hartwell and Paul Nurse,” Stegmeier says...regulate cell division. And that really got me hooked on genetics.” As a postdoc in Stephen Elledge at Harvard...a great chemistry teacher in 10th grade and then fell in love with science,” she says. In 11th grade she...high-resolution tracking of more than 1 million cancer cells under drug treatment.” Stegmeier especially finds...Belgium. “That’s where I not only got incredibly compelled by wet bench molecular research, but also became...these inefficiencies is pretty remarkable. I was compelled to be part of that.” He’s deposited AAV constructs...

  36. Tetbow: Bright Multicolor Labeling for Neuronal Tracing

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    ...imaging with single-cell resolution using chemical cocktails and computational analysis." Cell 157.3 (2014):... of the XFP genes is introduced at ~2 copies per cell per color, following a Poisson distribution. To ... optimal expression of XFPs. Any longer and the cells may start to show morphological abnormalities due...XFP genes have to be introduced at ~2 copies per cell per color to achieve the highest color variations... wait too long, as too much XFP is toxic to the cells. Inadequate expression levels of XFPs - Reduce the...video below depicting dendrite wiring of mitral cells.   Many thanks to our guest bloggers Richi Sakaguchi...circuitry with an index-optimized clearing agent." Cell reports14.11 (2016): 2718-2732. PubMed PMID: 26972009...

  37. Hot Plasmids: FRET-Based Biosensors

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    ... there is one problem: how to get that into the cells of interest,” Griesbeck said. Because they are chemical...understand the connection between molecular- and cellular-level events and behavior. It’s a problem that...advantage for observing events taking place inside cells that are on the move, such as T lymphocytes. Griesbeck...-binding protein, troponin C (TnC), from muscle cells. They identified a toadfish TnC variant which had...cAMP-responsive element-binding protein (CREB) in living cells. “Now we are making this really good,” Griesbeck...FRET-based sensor proteins, particularly for the intracellular imaging of transition metal ions, such as zinc...also present challenges in that they are found in cells at much lower concentrations than calcium, requiring...

  38. Plasmids 101: NGS Quality Control for Pooled Libraries

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    ...recently re-amplified our distribution stock of the Brunello Human gRNA library, and we thought it would be... Typically, Addgene uses electrocompetent Stbl4 cells when performing amplifications, and we use electroporation...DNA. This bias can be counteracted by growing the cells on solid plates, where each transformant forms its...own colony, uninfluenced by the growth of other cells. For libraries where the plasmids are all very similar...The colonies are then scraped off the plates and pelleted for DNA extraction. Prepare library product for...PCR. A diagram of the product we created for the Brunello library is shown in Figure 2. You can use a small...plotted as a histogram, the counts will give you a bell curve of the abundance of each component of your...

  39. Plasmids 101: Dimers and Multimers

    Type
    Blog Post
    ...at lower copy numbers within bacterial cells, and bacterial cells with multimers grow more slowly compared...remain intact, plasmid multimers function just as well as monomers. However, due to their larger size, ...stable and more prone to being lost in bacterial cells compared to monomers, although this can be prevented...increase the frequency of base substitution in human cells. J Biol Chem. 2011 Mar 25;286(12):10017-26. doi:...plasmids leading to the formation of plasmid multimers. Cell. 1976 Dec;9(4 PT 2):707-16. doi: 10.1016/0092-8674...supercoiled plasmid DNA concatemers in mammalian cells. J Gene Med. 2009 May;11(5):444-53. doi: 10.1002...essential for plasmid monomerization and stability. Cell. 1984 Apr;36(4):1097-103. doi: 10.1016/0092-8674...

  40. Building and Validating Recombinant Antibodies: A Case Study

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    Blog Post
    ...cryopreserved hybridomas, immortalized cells created by fusing an activated B cell that produces the antibody of...of interest with a myeloma cell. This RNA was then used to generate cDNA, which  was used as a template...of hybridoma generated mAbs is that if the frozen cell line becomes non-recoverable, the stock of the mAb...hybridoma stocks, without having to grow up the cells. This RNA could then be used to generate functional...immunofluorescence-immunocytochemistry (IF-ICC.) This method was chosen because cells transfected with a plasmid to express the target... could be screened in a high-throughput manner. Cells were labeled with both a hybridoma generated mAb...hybridoma mAb.  While screening by IF-ICC worked well for identification of the correct clones, it was...
Showing: 521 - 560 of 1028 results