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We narrowed to 297 results for: hal.2

Showing: 101 - 120 of 297 results

  1. Fluorescent Proteins 101: Aptamer Fluorophores

    Type
    Blog Post
    ... to DFHBI in the presence of that metabolite (Fig.2). Spinach eventually evolved into Spinach2, a more...PMID: 25713381. PubMed Central PMCID: PMC4364232. 2. Baker, Monya. “RNA imaging in situ”. Nature Methods...emerging environmental pollutants.” Front Chem. 2014; 2: 41. PubMed PMID: 25019067. PubMed Central PMCID: ...biosensors.” TrAC Trends in Analytical Chemistry 27(2):108-117 · February 2008. 11. Svensen, Nina and Jaffrey...technology has been studied and used in assays for over half a century; the Addgene blog has an excellent primer...

  2. Crowdfight, a Platform to Boost Scientific Collaboration During COVID-19 and Beyond

    Type
    Blog Post
    ...mammal called alpaca could neutralize the SARS-CoV-2 virus. But while the early results were promising,...receiving dozens of requests every day.    Figure 2: When a researcher needs a collaborator for a specific...Crowdfight will: 1) Find an expert with the right skill; 2) Confirm the expert’s suitability and availability...lockdown forced us to put our research projects to a halt. None of us works directly with viruses or transmissible...

  3. Predicting Adverse Reactions to Monoclonal Antibody Drugs

    Type
    Blog Post
    ...various components of the immune system. Figure 2: In-vitro models for immunogenicity prediction of ...drug reaction. This system would somewhat act like a 2-step verification. All approaches have their own limitations...Future Enhancements. Frontiers in drug discovery, 2, 952326. https://doi.org/10.3389/fddsv.2022.952326Joubert...Formation in Oncology: Clinical Relevance and Challenges. The oncologist, 21(10), 1260–1268. https://doi.org...

  4. Technical Design of a Western Blot

    Type
    Blog Post
    .... The most common options are 4 °C overnight or 1–2 hours at room temperature (RT). Most people choose...overnight at 4 °C and their secondary antibody for 1–2 hrs at room temperature, which allows them to reduce...Tris-glycine 6–400 kDa Tris-glycine 100 V, 1–2 hours Easy and cheap to handcast Short shelf-life...–40 kDa Tris-tricine 30 V, 1 hour or 100 V, 1–2 hours Good separation, quality, and stability ...separation of proteins of different sizes.  Table 2: Recommended gel percentages for various protein sizes...are consistent and stable at room temperature for 1–2 years and are available in formulas specific to your...response over a broad dynamic range that typically spans 2–5 orders of magnitude, which includes a wide dynamic...

  5. Droplet Digital PCR for AAV Quantitation

    Type
    Blog Post
    ... qPCR is that the results can vary by a factor of 2. This means that if you set up two identical assays...amplitude measured for each droplet in each well (Figure 2). A clean ddPCR should have a clear separation between...as low as 1C/s. Increasing the elongation time to 2 minutes and the denaturation time to 1 minute has ...generating the standard curve for qPCR can be challenging to get right. Here at Addgene, we use a linearized...droplet digital PCR for vaccine evaluation in challenged monkeys and TB diagnosis." Emerging microbes ...

  6. Plasmids 101: Protein tags

    Type
    Blog Post
    ...(you generally don’t want more tag than protein), 2-3 tags is increasingly common. Tandem affinity purification...TEV cleavage site (more on that in a moment), and 2 ProtA IgG-binding domains. TAP has since come to encompass...found in this paper (available at Addgene).   Table 2: Protease recognition sites commonly used with tags...purify proteins can face any number of technical challenges depending on their protein of interest. After...After several decades of trying to address these challenges, researchers have amassed a considerable molecular...

  7. Visualizing Translation at the Single Molecule Level

    Type
    Blog Post
    ...Find ribosome profiling plasmids at Addgene. 2. Halstead, James M., et al. “Translation. An RNA biosensor...the reporter mRNA is labeled by PCP-mCherry (Figure 2). The 3’ UTR also contains a CAAX sequence to bind...translation inhibitors cycloheximide and puromycin, Halstead et al. verified that translation is necessary ...translation is known to be important in development, and Halstead et al. used TRICK to examine translation of Drosophila...

  8. Isolating a Monoclonal Cell Population by Limiting Dilution

    Type
    Protocol
    ...Day 1: Seed individual cells in a 96-well plate Day 2–14: Monitor cells for growth and expand cells Day ...Biological Safety Cabinet 0.5–10 µL single channel pipette 2–20 µL single channel pipette 20–200 µL single channel...channel pipette 200–1000 µL single channel pipette CO 2 incubator Pipet controller Hazardous waste container...approximately 50–60% confluent. For 293T cells this is about 2 × 10 6 cells in a 10 cm dish. Each 10 cm dish should...the highest or lowest transgene expression ( Figure 2 ). Sample Data Figure 1: Generation of monoclonal ...inferred by the differences in colony size. Figure 2: Cas9 expression in monoclonal cell lines generated...medium at a reduced antibiotic concentration (e.g., half the concentration that was used for the initial ...

  10. Antibodies 101: Avoiding the Mouse on Mouse Mess in IHC

    Type
    Blog Post
    ...recognize Fab fragments from the same species and 2) these fragments will be more likely to bind (and ...epitopes recognized by the secondary.   Figure 2: There are a few common solutions to avoid background...staining can be a challenge in the best of cases. But it can be particularly challenging when your only ...

  11. Important Considerations in Optogenetics Behavioral Experiments

    Type
    Blog Post
    ... will repeatedly nose-poke at the active hole [1, 2].    Place preference/place aversion also models reinforcement...spend most of its times in the opposite chamber) [2]. Laser activation is usually controlled automatically...two different ways 1) your hypothesis is wrong or 2) there is some problem with the technical implementation...PMID: 21716290. Pubmed Central PMCID: PMC3775282. 2. McDevitt, Ross A., et al. "Serotonergic versus nonserotonergic...than ICSS. The test chamber is divided into two halves that are distinguished visually through different...

  12. RNA Interference in Plant Biology: New Tools for an Old Favorite

    Type
    Blog Post
    ... analogs such as 2-fluoroadenine (2-FA), organisms with functional APT will convert 2-FA to cytotoxic ...selection of plants undergoing RNAi, and LIIbeta F 1-2 RNAi, which allows assembly of intron-spliced hpRNAs...nucleotides, resulting in death. In the presence of 2-FA, potent gene silencing of APT is required for survival...such as in the model flowering plant Arabidopsis thaliana (Li et al., 2013; Zhang et al., 2018) and the ...

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

    Type
    Blog Post
    ... you can read more about them in review articles (2–4). You can also take a look at my recent paper to...Development 118, 401–15 (1993). PubMed PMID: 8223268. 2. J. B. Duffy, GAL4 System in Drosophila : A Fly Geneticist...two genes by one Gal4 driver will produce roughly half as much protein as overexpression of each gene individually...384–391 (2004). PubMed PMID: 15262411. 4. T. D. Southall, D. A. Elliott, A. H. Brand, The GAL4 System :...Marka, J. C. Kagan, Environmental Stress Causes Lethal Neuro-Trauma during Asymptomatic Viral Infections...

  14. Targeted Mutagenesis with EvolvR

    Type
    Blog Post
    ...while increasing the standard mutation rate by only ~2-fold over nCas9 levels. Two sets of modifications ...10-3 mutations per nucleotide per generation, and (2) the addition of the thioredoxin-binding domain (TBD...components of EvolvR and its potential applications. Challenges of existing mutagenesis methods Several continuous...drawbacks with these approaches that makes it challenging for users to achieve targeted diversification...PMID: 30253789. PubMed Central PMCID: PMC6154929. Halperin, Shakked O., et al. "CRISPR-guided DNA polymerases...

  15. Treating Muscular Dystrophy with CRISPR Gene Editing

    Type
    Blog Post
    ...nonsense-mediated decay. Although Nelson et al. observed only 2% genome editing in one experiment, they found the ... cells throughout the body, especially stem cells 2. Remove any immunogenicity of the AAV vector Once ...Science. 2015 Dec 31. doi:10.1126/science.aae0169 2. Long C, Amoasii L, Mireault AA, McAnally JR, Li H...Precision genome editing faces the challenges above and more. One chief challenge is upregulating HDR in mature...complex oligonucleotide treatment comes with many challenges, researchers have begun to explore genome editing... the time to clinical approval. Even with the challenges precision editing faces, it’s encouraging to ...WX, Scott DA, Gootenberg JS, Kriz AJ, Zetsche B, Shalem O, Wu X, Makarova KS, Koonin EV, Sharp PA, Zhang...

  16. CRISPR 101: Epigenetics and Editing the Epigenome

    Type
    Blog Post
    ...form or as ready-to-use lentivirus. Figure 2: dCas9-Tet1 demethylates cytosines at promoters and...pX330a dCas9-LSD1 for targeted removal of H3K4me1/2 and H3K9me2 histone modifications. Like the TALE-LSD1...approach for targeted DNA demethylation. Cell Discovery 2: . https://doi.org/10.1038/celldisc.2016.9 Find plasmids... demethylase to demethylate enhancer regions (Mendenhall et al., 2013). By comparing gene activation when...sorting and selection of transduced cells. Grant Challen’s lab also created constitutive (pCMV-dCas9-D3A)...contributed to updating this article. References Haldeman JM, Conway AE, Arlotto ME, et al (2018) Creation...Fowler T, Rees R, Kramer A, Martens A, Edwards JR, Challen GA (2016) Reprogrammable CRISPR/Cas9-based system...

  17. CRISPR Methods for Bacteria: Genome Engineering, CRISPRa, CRISPRi, Base Editing, and More

    Type
    Blog Post
    ... pCRISPomyces-2 contains Cas9 and a gRNA cassette. The simpler system of pCRISPomyces-2 displays a higher...and a subsequent curing protocol that requires only 2-3 hours incubation. E. coli and T. citrea scarless...four major components including (1) a CRISPR RNA, (2) four proteins forming the QCascade DNA-targeting ...DSB) are lethal. Addgene depositor Luciano Marraffini’s lab took advantage of this lethality to design...organisms, CRISPR-induced double stranded breaks are lethal because the non-homologous end-joining (NHEJ) repair...Cas9 will cleave the gene of interest, inducing a lethal DSB.     This system is distinct from those used...While the activation of transcription is a more challenging problem than repression, scientists have deposited...

  18. Antibodies 101: Reading a Flow Plot

    Type
    Blog Post
    ...A-B+ cells. If you look at the dot plot in Figure 2, you might assume that all cell populations are positive... the different populations. In the case of Figure 2, one population is B- and another two are B+. Our ...are in the lower right corner.      Figure 2: This flow plot has two populations that are B+ and...from flow cytometry experiments can be a little challenging to understand if you’ve never done any flow. ...., Sait, K., Anfinan, N., Al-Wasiyah,  mohammad khalid, Huwait, E., Gari, M., & Al-Qahtani, M. (2018)....

  19. Adenovirus Guide

    Type
    Guide
    ...use of pAdEasy-2 (Addgene #16401) can increase the capacity of the rAdV vector. pAdEasy-2 does not contain... incorporate additional genomic deletions. Figure 2: First-generation rAdV vectors. Created with BioRender.com...and have to be handled at biosafety level two (BSL-2). They are highly immunogenic and can trigger a strong...era of personalized medicine . Genes & Diseases, 4 (2), 43–63. https://doi.org/10.1016/j.gendis.2017.04.001...adenoviruses using the AdEasy system . Nature Protocols, 2 (5), 1236–1247. https://doi.org/10.1038/nprot.2007.135...Adenoviral vector vaccine platforms in the SARS-CoV-2 pandemic . NPJ Vaccines, 6 (1), 97. https://doi.org...Adenoviral backbone plasmid that lacks E1 and E3. pAdEasy-2 Adenoviral backbone plasmid that lacks E1, E3, and...

  20. Pathways Over Time Plasmids Engage Students in Functional Genomics Research

    Type
    Blog Post
    ... undergraduate students (1, 2), but educators face significant challenges in designing suitable projects... by western blotting in other experiments (Figure 2).   Complementation reveals conservation of function...Biology Education: A Call to Action, Washington, DC. 2. President's Council of Advisors on Science and Technology...
Showing: 101 - 120 of 297 results