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Showing: 451 - 500 of 929 results

  1. Plasmids 101: Positive and Negative Selection for Plasmid Cloning

    Type
    Blog Post
    ...synthetize all compounds needed for growth, or auxotrophy, the inability to synthesize a compound needed to...an essential amino acid), and a plasmid that complements the auxotrophy (i.e., synthesizes the critical...yeast prototrophy kit containing 23 plasmids to complement auxotrophy. Negative selection: Cells that have...

  2. An Addgene Summer Internship

    Type
    Blog Post
    ... had to get used to the workspaces, as well as complete the lab safety and cybersecurity training before...project is a major project given to each intern to complete and present on.    Figure 1: Addgene's...size should be a max of 500 bp (preferably). I compiled a list of three options from the Addgene catalog...Toj-Villagran is a rising senior in high school who just completed their summer internship at Addgene. They plan ...

  3. CRISPR 101: Cas9 Nickase Design and Homology Directed Repair

    Type
    Blog Post
    ...question is: how should the gRNAs be oriented in comparison to each other? The gRNAs must target different...different strands to create a DSB, but this can be accomplished with either a PAM-in or PAM-out orientation....preferences using HEK293 cells. They started by comparing D10A and H840A total editing efficiency in PAM-in...donor with 40 bp homology arms. The HDR efficiency compared favorably to using WT Cas9 with the left gRNA ...

  4. An Addgene Co-op: The Intersection of Fun, Friendship, and Work

    Type
    Blog Post
    ... to complete a six month work experience called a co-op to receive a degree. Northeastern compiles a list...Thankfully, my managers set aside time for me to complete an online social media certification on social...petting dogs, and winning the office Just Dance competition.    Special thanks to my manager, Sr. Marketing...

  5. Addgene-Seeding Labs Plasmid Grant: Accelerating Science Globally - 5 Scientists, 4 Countries & 3 Continents!

    Type
    Blog Post
    ...projects spanning from the identification of plant compounds and insecticidal proteins to the development of... reside within living plants and that produce compounds with various antimicrobial activities (1). She... newly identified proteins and enzymes. These compounds will later get tested for their potential agricultural...alternative source of essential omega-3 fatty acids complementing vegetable oil, a somewhat poor natural source...

  6. Tips for Improving Your Next Manuscript

    Type
    Blog Post
    ...t guarantee a publication. There is an art to composing manuscripts and the ASM Scientific Writing and...writer, I was unsure if I should possibly take a composition class in the English Department. He suggested... coming to the writing workshop in D.C. had to compose and submit a manuscript that they were working ...portion of the paper will be generated. After you complete a publication quality figure, produce the figure...

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

    Type
    Blog Post
    ...CRISPR screen data are also available via comprehensive pre-compiled downloads and via a full-featured REST... to stably archived large-scale datasets by computational biologists. Figure 1: The BioGRID Open... from humans and other species. By accurately compiling genome-wide datasets, which are often obscured...

  8. Viral Genome Sequencing - A One Stop Shop for AAV Identity and Serotype Confirmation

    Type
    Blog Post
    ...platform that allows us to provide scientists with complete plasmid sequences. After launching our viral service...present SMRT sequencing only works for self-complementary AAVs and the TA-based ligation and tagmentation...molecules in the DNA extract associate with complementary strands, enough double-stranded species exist...adeno-associated virus particles containing complementary polynucleotide chains. J Virol 9(2): 394-396...

  9. Summer SciComm Series: Master of SciComm

    Type
    Blog Post
    ...and not solely to ensure a profit. Often, these companies believe they possess something that will truly...the public’s perception of head injury risk. In completing a master’s thesis in science communication, I... I had the chance to carry out a comprehensive review of the literature informing the notion of “heading... Department of Science Communication where he completed a master's degree in 2020, his research explores...

  10. Plasmids 101: TOPO Cloning

    Type
    Blog Post
    ...The technique relies on the basic ability of complementary basepairs adenine (A) and thymine (T) to hybridize...deoxyadenosine (A) at the 3' ends of PCR products. The complimentary “T” in the pair comes from a topoisomerase I-linearized...overnight. 4. Transform TOPO Cloning Reaction into Competent Cells: You can use your standard lab protocol ...transforming your reaction into chemically or electro-competent E. coli - excess salt causes arcing during electroporation...

  11. Behind-the-scenes of the Isolation of the Thermostable IgnaviCas9 From a Yellowstone Hot Spring

    Type
    Blog Post
    ...volunteered to do the sampling and I happily agreed to accompany him to YNP the following year. Geoff Schiebinger...microfluidic parallelization to sequence this highly complex hot spring sample. The sample was separated into...cells each for sequencing, allowing for lower complexity with higher throughput and rigorous assembly....ThermoCas9 from the van der Oost lab. Phylogenetic comparison indicates that like GeoCas9 and ThermoCas9, IgnaviCas9...

  12. Extracurricular Activities for a Strong Science Career Path

    Type
    Blog Post
    ...great approach to fully understand and acquire competitive skill sets in a myriad of professions is to engage...Mentorship Leadership Science writing competitions (e.g. Access to Understanding)   Writing...Communication Networking Publicity Business competitions   Leadership Management Marketing ...opportunity to tailor their career paths for competitive positions both within and outside academia. Use...

  13. Antibody Validation for Flow Cytometry

    Type
    Blog Post
    ...intracellular labeling because the intracellular compartment may contain cross-reacting antigens not present...difference in the efficacy of your antibody. When compared, 4% PFA and 0.1% saponin; 4% PFA and 0.1% Triton...intensity in the KO population (pink histogram) compared to the WT cells (green histogram). The unfilled..., like RNAseq or proteomics, it is possible to compare antibody labeling of different cell types within...expression of the target. In this case, you could compare the antibody’s labeling of the cell lines to the...

  14. Plasmids 101: Gram Negative and Positive Bacteria

    Type
    Blog Post
    ... bacteria is due to differences in cell wall composition. Cell wall structure Gram positive bacteria, ...is added, it interacts with the CV+, creating a complex within the peptidoglycan layer, staining the cell...the peptidoglycan layer, trapping the large dye complex created by the interaction of CV+ and I- in the...type of bacteria it is being used in, and how components of the plasmid should change based on bacteria...

  15. Using CRISPR/Cas9 to Edit Disease Out of the Genome

    Type
    Blog Post
    ... bulk of which you can find below. CRISPR for complex genetic diseases As CRISPR technology becomes increasingly...using it to investigate potential therapeutics for complex genetic diseases. So we’re revisiting this post...look at CRISPR’s potential for use in treating a complex genetic diseases, Alzhiemer’s disease (AD).  Troy...fibrosis (8).   The difference with CRISPR/Cas9 in comparison to other genome editing techniques is its ease...

  16. Creating accessible biology activities in schools with BioBits

    Type
    Blog Post
    ...cellular level. The reason for this is cost and complexity – anyone who works in the lab knows how expensive...expensive tiny amounts of reagents are, how complicated (and again, expensive) the different machines are...kits based on freeze-dried, cell-free (FD-CF) components. Making proteins outside of a cell - cell free..., feel free to check out one of our kits and accompanying curriculum to see if any of the activities fit...

  17. Visualizing Translation at the Single Molecule Level

    Type
    Blog Post
    ...time. Both are helping researchers explore the complexity of translational control in cellular physiology... processing to avoid disrupting ribosome-mRNA complexes. Recent work has adapted this technique to monitor...color switching, TRICK requires the following components: a GFP protein that binds to the coding region... with mRNAs usually translated in a specific compartment. Despite these limitations, these techniques ...

  18. How Dry Ice Affects Viral Vector Sample pH & How to Avoid It

    Type
    Blog Post
    ...liquid solution and pH paper to measure the pH. We compared the dry ice-stored samples to controls that were...ice for 72 hours and then vented the tubes by completely removing the caps, incubating the tubes in an...and then recapping the tubes prior to thaw. We compared vented and non-vented tubes to controls that were...various recovery periods between 0 and 96 hours. We compared these samples to controls that were only stored...

  19. Addgene Depositors Get More Citations

    Type
    Blog Post
    ...plasmids as if they comprised 10 percent. Based on this new re-weighting, we can compare citations where ...Pamela McA'Nulty and Zachary Tribbett for helping compile the data used in this work. Samantha Zyontz is... topics including biotech and research tools, computing performance, and randomized control trials in ...

  20. Hot Plasmids - December 2019 - CRISPR pooled library, LipoGlo, CRISPRa, and Voltage Imaging Tools

    Type
    Blog Post
    ... the vascular wall. ApoB, the main structural component of atherogenic lipoproteins, shuttles lipoproteins...Thus, the bEFPs bind the UAS via the dCas9-gRNA complex which allows transcription from the promoter.  ...should serve as both a powerful tool for generating complex regulatory systems and further the study of natural...hypothesizes that this may be due to different lipid compositions of C. elegans membranes, which might beneficially...

  21. Optimizing Donor DNA for Enhanced CRISPR Genome Editing

    Type
    Blog Post
    ...strand in Figure 1 below) was free to anneal to complementary ssDNA molecules in vitro. Figure 1: Cas9...discovered that single stranded donor DNA: Complementary to the released strand, 127bp in length, With...target DNA that was accessible to anneal with complementary single stranded DNA. We thus wondered if such... any of the error-prone repair that normally accompanies Cas9 cutting. We still don’t know the mechanism...

  22. Addgene Welcomes John Doench to the Advisory Board

    Type
    Blog Post
    ...Sharp. After a few rotations, Doench decided to complete his PhD work in Sharp’s lab.  Doench says, “what...what I loved about [RNAi], ... it was just this complete black box. The RNAi process had just been discovered...evolved into the Genetic Perturbation Platform and encompasses a variety of technologies to functionally investigate...he says, “In the case of CRISPR, the hype was completely justified… it has been just an incredibly useful...

  23. Fluorescent Proteins 101: Luciferase

    Type
    Blog Post
    ...because of the nature of the chemical reaction compared to using a high-intensity laser to rapidly excite...not required to initiate the reaction. Table 1 compares some general advantages and disadvantages of luminescence... Depending on the specific luciferase used, a compatible substrate must be provided for the enzyme to ... will result in luciferase expression and more complex cell treatments can be designed to further explore...

  24. Finding and Joining Your Dream Lab

    Type
    Blog Post
    ...guide for the quest of a perfect "fit". How competitive is the lab? Some labs have only one author plus...labs around the world. You may prefer a more competitive style to spur you to succeed or you might enjoy...enjoy working closely with others to accomplish your goals. You must understand your own style before making...Finding your dream lab While none of us will feel completely happy at work all of the time, the goal is to...

  25. Synthesized by Ginkgo Bioworks, Shared by Addgene: SARS-CoV-2 Plasmids for Many Expression Systems

    Type
    Blog Post
    ...available. Plasmids are continually added as they complete QC and you can find all available plasmids listed...proteins, and membrane proteins. Although all viral components are available in bacterial expression plasmids...batches of samples. As soon as that process is complete, QC scientists prepare Sanger reactions by mixing...are met, we are taking a unique hybrid approach compared to our standard pipeline. For the first ~50 plasmids...

  26. Targeted Mutagenesis with EvolvR

    Type
    Blog Post
    ...library used. EvolvR: key components The EvolvR system has two key components: the error prone PolI3M that...variants in bacteria. Read on to learn about the key components of EvolvR and its potential applications. Challenges...interest. A gRNA is used to direct the PolI3M-nCas9 complex to a DNA site of interest, which nCas9 nicks and...

  27. Hot Plasmids December 2018 - C. elegans RNAi, TurboID, Detecting Viral Outbreaks, and Protein Binding Specificity Tools

    Type
    Blog Post
    ...to physically set up in the lab, RNAi is often completely ineffective or results in low penetrant phenotypes...membranes, organelles, and other subcellular compartments is via proteomics--the study of the distribution...low-cost, cell-free detection assay that can be completed without special equipment. What’s more, detection...encodes the LacZ𝛼 protein. Once expressed, LacZ𝛼 complexes with LacZ𝜔 and forms an active LacZ tetramer....

  28. Why and how to start studying chromatin (with Addgene's help)

    Type
    Blog Post
    ...being depicted, we can’t forget that those other components are still there in real life.  Certainly referring...interact with each other. Chromatin regulation is complex and influences--directly or indirectly--many other...PIs, articles, genes, and blog posts are not comprehensive, so just because you do not see a name or gene...phosphoinositol-dependent binding of the SWI/SNF-like BAF complex to chromatin after T lymphocyte receptor signaling...

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

    Type
    Blog Post
    ...Konermann et al. Plasmids here How do Cas13d enzymes compare to other Cas13s? Like other Cas13 enzymes, the ...cleavage pattern is necessary. Konermann et al. compared CasRx to multiple RNA regulating methods: small... the clear winner with median knockdown of 96% compared to 65% for shRNA, 53% for CRISPRi, and 66-80% ...Addgene. Yan, Winston X., et al. “Cas13d Is a Compact RNA-Targeting Type VI CRISPR Effector Positively...

  30. Quick Guide to Working with Drosophila Part 3: Genome Engineering in Flies

    Type
    Blog Post
    ...chromosome. Targeted Insertion allows for direct comparisons between lines expressing different variants of...chromosome your transgene has integrated. This can be accomplished with a few matings to balancer chromosome lines...into a plasmid, you can send your plasmid to a company. For a few hundred dollars, they will inject the... your strategy and the services offered by the company, you might have to map into which chromosome the...

  31. Opto-Nanobodies: Using Light to Manipulate Cell Signaling and Protein Purification

    Type
    Blog Post
    ...controlling cell signaling. Key components of OptoNBs OptoNBs have two key components: an AsLOV2 domain and a ...activity in cells.  Let’s shed some light on the key components of OptoNBs, how they are generated, and their...variable heavy chain antibody domain.  How do these components interact? When fused together to form an OptoNB...

  32. REPLACR Mutagenesis: Replacing In Vitro Recombination Methods

    Type
    Blog Post
    ...No No Yes Yes Lab Made Competent Cells Required Yes No No No Comparison of REPLACR-mutagenesis with...mutations, large deletions/insertions require complicated, often costly in vitro assembly methods. A new...-mutagenesis also requires making your own electrocompetent cells in which the phage recombination machinery...

  33. Antibodies 101: Affinity Reagents

    Type
    Blog Post
    ...camelid antibodies to antibody fragments to tools completely unrelated to antibodies.  Why use an alternative...entirely new affinity reagent.    Fig. 1: A comparison of an antibody (left) to several alternative ...through engineering instead of cleavage. They are comprised of the variable heavy and light chain regions,...immunogenic, but they have lower affinity and longevity compared to antibodies, along with a higher likelihood ...

  34. Antibodies 101: Polyclonal Antibodies

    Type
    Blog Post
    ... the bulk purification steps keep the cost low compared to other antibody formats. This low barrier to...is no more of that exact polyclonal antibody composition available. If you’re going to need a large quantity...antibodies produce a higher level of background noise compared to monoclonal or recombinant antibodies. This ...polyclonal antibodies for your experiment and how they compare to other antibody options out there, you’re set...

  35. dTAG - You're it!

    Type
    Blog Post
    ...degradation In the Bradner and Gray labs, we synthesize compounds that enable selective removal of proteins-of-interest...FKBP12F36V-tagged proteins. The dTAG molecule induces ternary complex formation between the FKBP12F36V-fusion and the... for degrading targets in different cellular compartments, and in vivo utility. dTAG molecules will be...lentiviral plasmids can serve as a rapid means of comparing functionality of N- and C-terminal fusions and...

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

    Type
    Blog Post
    ...is dependent on proteasome function. TRIM-away compared favorably to drug inhibition, with TRIM-away targeting...larger cell populations, you can deliver these components together using electroporation. In these bulk...250 nm), and studying endocytosis is further complicated by the reliance on pH-based biosensors used to...rapid and reversible recruitment of chromatin complexes to any genomic locus in any cell type. By varying...

  37. RANbodies: Reporter Nanobody Fusions

    Type
    Blog Post
    ...process that’s difficult for individual labs to complete. The nanobody based RANbody platform from the ...the sample due to the smaller size of RANbodies compared to antibodies. Staining with multiple HRP-RANbodies...sample is stripped. HRP-RANbodies are, however, compatible with most multi-labeling immunohistochemical ...antigen with a primary/ labeled secondary antibody. Compatible with multi-labeling protocols when other antigens...

  38. Oh, The Places You Can Go: Careers in Science Communication - Writing for a Research Institute

    Type
    Blog Post
    ...show. After graduating with a master’s, Susan completed a science writing internship with the National...planning.” To beef up her resume, she decided to complete a certificate in Strategic Communications and ...the most satisfying parts of the job is having completed one of these difficult pieces: “It’s probably ...science communication] needs. I worked for one company, but there were a ton of others that also did work...

  39. All in a Twist: dsRNA

    Type
    Blog Post
    ...silencing complexes (RISCs), which detect dsRNA in the cytoplasm and use it as a guide to complementary RNA ...RNA sequence. Once a compatible sequence is found (typically RNA that the RNAi is designed against), RISC...and will dissociate when gene transcription is complete. However, transcription of thousands of genes ...

  40. Antibodies 101: Conjugation

    Type
    Blog Post
    ... are considering to make sure that they’ll be compatible with your instruments, each other, and your target...target. I love FPbase’s Spectra Viewer tool for comparing different fluorophores. We also touched on fluorophores... sure to check that your antibody’s buffer is compatible with your conjugation kit, since many common ...reactions. If needed, you can often remove the incompatible agents by dialysis or gel filtration.    There...

  41. Plasmids 101: Cre-lox

    Type
    Blog Post
    ...recombination events. The system consists of two components derived from the P1 bacteriophage: the Cre recombinase...recognition site. The P1 bacteriophage uses these components as part of its natural viral life-cycle, and ...and researchers have adapted the components for use in genome manipulation. Cre recombinase, originally...other sites of the same type, but are not cross-compatible. Using different lox site variants allows Cre...

  42. A Guide to Designing a Scientific Poster: Content, Layout, and More

    Type
    Blog Post
    ...audience’s eye and draw people in. That can be accomplished with an exciting title and attractive figures...in less than a couple of minutes. How does one accomplish all of that? What follows are some tips to help...logical progression of your work, and you can accomplish this by employing a neat and organized format...want to make sure that the color of your font is compatible. Use high-contrast between these colors so that...

  43. Antibodies 101: Stripping and Reprobing Western Blots

    Type
    Blog Post
    ...’ll share the basic composition of these solutions below. Alternatively, companies including ThermoFisher... tool to identify a protein of interest in a complicated solution like cell lysate. But they can be a ...lost each time you strip a blot, you should not compare the strength of bands from different rounds of ...

  44. Overcoming the Challenges of Lentiviral Production

    Type
    Blog Post
    ... formation of replication competent lentivirus (RCL). Replication competence can arise from recombination...drive high levels of expression of certain viral components; according the manufacturer these changes result...downstream. When assessing lentiviral titer, don't compare apples to oranges After choosing a packaging system...

  45. Rabies and Neuronal Tracing

    Type
    Blog Post
    ...optogenetic or chemogenetic receptors) enables complex control of specifically defined neuronal populations...how newly generated neurons integrate into the complex pre-existing circuit of the brain. Another study...anatomical regions of the brain receive inputs and compute outputs. In general, RABV monosynaptic tracing ...circuitry, and continue to contribute answers to the complex questions of how the brain operates. References...

  46. Viral Vectors 101: Chemogenetics

    Type
    Blog Post
    ... is important to fully characterize the compound, or compounds, you choose to ensure reliable results ...In order to understand the complex wiring of the brain and the underpinnings of neurological disease,...pharmacodynamic actions of clozapine-N-oxide, clozapine, and compound 21 in DREADD-based chemogenetics in mice. Scientific...

  47. Adapting Toehold Switches to Detect Zika Virus

    Type
    Blog Post
    ...up clear tasks for each member of the team to accomplish so that many elements of the project were able...involved definitely contributed to our ability to complete the project on such a short timescale. Our article...extensive infrastructure of molecular and electronic companion technologies that has transformed the assay’s ...of Zika Virus Using Programmable Biomolecular Components." Cell (2016). PubMed PMID: 27160350. Find Plasmids...

  48. Adenoviral Delivery of CRISPR/Cas9 Aims to Expand Genome Editing to Primary Cells

    Type
    Blog Post
    ...protein capsid encapsulating the genome is majorly composed of three proteins that can be easily modified ...Delivering gRNA and Cas9 using adenoviral vectors As compared to other viral vectors, AdVs can provide a large...AdV-mediated transduction of gRNA:Cas9 ribonucleoprotein complexes into transformed and non-transformed cells yielded...vector delivery of RNA-guided CRISPR/Cas9 nuclease complexes induces targeted mutagenesis in a diverse array...

  49. Fluorescent Proteins 101: Fluorescent Protein Timers

    Type
    Blog Post
    ...better at producing a compound of interest, thus allowing them to optimize the compound production process...fluorescent proteins have usurped old methods or complemented them in ways that make them much easier. A special...Although it is possible to make a timer system composed of two separate FPs (one that matures quickly ...

  50. Light Sheet Fluorescence Microscopy

    Type
    Blog Post
    ... capture large field-of-views (6). For more comprehensive reviews on several other LSFM configurations...multiplying couple-charged device (EMCCD) or complementary metal-oxide semiconductor (CMOS) sensors, it...labeling in whole tissue (15).  CLARITY is also compatible with LSFM, a method termed COLM (clarity-optimized...spatiotemporal control over labeling methods as compared to transgenic animals (17).  These viral plasmids...
Showing: 451 - 500 of 929 results