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We narrowed to 490 results for: ARA-2

Showing: 61 - 80 of 490 results

  1. Fluorescent Protein Guide: Subcellular Localization

    Type
    Collection
    ...sequence YFP Thorben Dammeyer 54520 mCherry-Peroxisomes-2 Peroxisome Peroximal Targeting Signal 1 (PTS1) mCherry...Centrin2 Centrioles (dependent on cell cycle) Centrin-2 EGFP Erich Nigg 41151 pEGFP Cep170 C-term Centrioles...GolgiCFP Golgi apparatus eNOS(1-33) CFP Alexandra Newton 36205 pmTurquoise2-Golgi Golgi apparatus B4GALT1(1...10 Golgi apparatus MannII mGold Francois St-Pierre 85050 pmScarlet-i_Giantin_C1 Golgi apparatus Giantin...pmScarlet-H_Giantin_C1 Golgi apparatus Giantin mScarlet-H Dorus Gadella 85048 pmScarlet_Giantin_C1 Golgi apparatus Giantin ...Giantin Golgi apparatus Giantin mTurquoise2 Dorus Gadella 98880 mNeonGreen-Giantin Golgi apparatus Giantin ...subcellular structures. To compare with a well-characterized marker's localization pattern, you should fuse...

  2. Antibody Neutralization Response Against Pseudoviruses Expressing SARS-CoV-2 Spike Protein Variants

    Type
    Blog Post
    ..., Schmidt AG, Iafrate AJ, Naranbhai V, Balazs AB (2021) Multiple SARS-CoV-2 variants escape neutralization...months, we’ve seen the emergence of several SARS-CoV-2 variants coincide with increased vaccination efforts...They were able to tweak this assay to study SARS-CoV-2 spike variants. “We created a series of spike expressing...relative ability to neutralize.”   Figure 2: Sera used in the study came from individuals who ...This sera was used in the high-throughput SARS-CoV-2 pseudovirus neutralization assay. Neutralization was...viruses are quite genetically distinct from SARS-CoV-2 and its variants. Yet, the B.1.351 variant, which ...effectiveness of current vaccines against SARS-CoV-2 variants, this study was the first to compare so many...

  3. Antibodies 101: Normalization and Loading Controls for Western Blots

    Type
    Blog Post
    ...the resulting fraction is reported. Figure 2: Normalization with loading controls. Under each lane...control band is weaker in condition 1 than condition 2, the normalized values for the two conditions are ...opposite change in your loading control. Assumption #2: The loading control can be accurately quantified ...significantly lighter in “condition 1” than “condition 2.” By eye, the loading control appears about the same...) are smaller than the analogous green “condition 2” points by the exact same value. For the two solid...consider a different loading control. Assumption #2: The loading control can be accurately quantified ...factors we’ve covered in our previous blog posts (1, 2) on western blotting. If the abundance of your POI...

  4. Viral Vectors 101: Virus Safety

    Type
    Blog Post
    ...cancer. These viruses need to be handled in a BSL-2 (or BSL-2+) lab and require additional safety measures.... BSL-1. In BSL-2 labs, eye protection is required any time you have to work with BSL-2 materials, such...respiratory diseases in humans, earning this virus a BSL-2 classification. Lentivirus (a type of retrovirus) ...recommended or required in BSL-1 and required in BSL-2 labs. When working with lentivirus, many organizations...sprayed into the air. For lentivirus and other BSL-2 materials, you’ll need to use aerosol-tight caps, ...percentage of the decontaminate to be used for BSL-2 waste, and of course, if you are working with a substance...transfer plasmid), the envelope (Env) gene on a separate plasmid (envelope plasmid), and the polymerase...

  5. Viral Vectors 101: Pseudotyping

    Type
    Blog Post
    ...safely. Recently, many SARS-CoV-2 researchers began pseudotyping the SARS-CoV-2 spike protein into a lower ...SARS-CoV-2 virus. Scientists have been using similar pseudotyping methods with other viruses in a BSL-2 lab... et a., 2018).   Figure 2: To pseudotype with the SARS-CoV-2 spike protein, scientists encode...production uses three plasmids: (1) The transfer plasmid, (2) the envelope plasmid, and (3) the packaging plasmid...example, production of vesicular stomatitis virus, HIV-2, and human foamy virus in cell lines expressing galactosyl...for Pseudotyping Lentiviral Particles with SARS-CoV-2 Spike Protein for Neutralization Assays. Viruses 12...Are Inactivated by Human Serum. Molecular Therapy 2:218–222 . https://doi.org/10.1006/mthe.2000.0116 Gutierrez-Guerrero...

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

    Type
    Blog Post
    ....9 (parental) 46 +/- 0.7% AAV.PHP.S 82 +/- 2% Table 2: Transduction efficiency of AAV.9 versus AAV.PHP.S...figure 2) to allow for efficient IV delivery of virus to the CNS or PNS (figure32, tables 1 & 2). These...because: 1) their genomes are easy to manipulate, 2) they have long-term expression; and 3) they have ... On average, AAV.PHP.eB and AAV.PHP.S had 1.5- to 2-fold higher GFP intensity compared to the strains ...stained for a neuronal marker). Check out tables 1 and 2 below for a summary of the quantified transduction...cells are labeled with their own unique color; and 2) low levels of labeling so that a cell’s projections...maximize the number of cells that are labeled. Part 2: An AAV expressing the inducer (i.e. tet-off transactivator...

  7. Lambda Red: A Homologous Recombination-based Technique for Genetic Engineering

    Type
    Blog Post
    ...generate 2 possible products: 1) a partially dsDNA duplex with single-stranded 3’ overhangs or 2) if the...recombineering system has three components (Figure 1): 1) Exo, 2) Beta, and 3) Gam. All three are required for recombineering...generic lambda red recombineering experiment (Figure 2). In the following sections, key steps that differ...confirmation of recombinant clones.    Figure 2: Overview of using Lambda Red recombineering system...pairs of homology to the targeted insert site (Figure 2, top).  These primers are typically ~70 nucleotides...bacteria that has key MMR proteins knocked out or 2) specially design ssDNA oligos to avoid MMR: 1) E....will have more unintended changes to their genomes. 2) Designing ssDNA oligos that avoid MMR activation:...

  8. Scientists Map the SARS-CoV-2-Human Interaction Network

    Type
    Blog Post
    ...comprehensive SARS-CoV-2 virus-human interaction network in which we expressed 26 of the 29 SARS-CoV-2 viral proteins...all became acutely aware of the threat of SARS-CoV-2. Like many in the science community, we’ve been motivated...identifying these potential human helpers of SARS-CoV-2 (especially the ones that are druggable) is a key ...just at the beginning of understanding how SARS-CoV-2 interacts with human proteins. For example, we used... joining in this worldwide fight against SARS-CoV-2. As we’ve all been affected in various ways by the...details. Our set of constructs comprises all SARS-CoV-2 viral proteins except for Nsp3 and Nsp16, and includes...is going on there!).  Find the Krogan Lab SARS-CoV-2 plasmids (Note: The Nsp6 construct is undergoing QC...

  9. Fluorescent Proteins: FRET

    Type
    Collection
    ...referred to as intermolecular or bimolecular FRET) or (2) conformational changes within a protein where the...protein-protein interactions where each protein is separately fused to a donor or acceptor molecule (also referred...pairs with mTurquoise2 Mastop et al. (2017) Characterization of a spectrally diverse set of fluorescent...

  10. Plasmid Preps: Different Purity, Different Quantities, Different Uses

    Type
    Blog Post
    ... be extracted from small bacterial cultures (i.e. 2-3 ml) with or without commercially available kits ...ratios measured at 260 and 280 nm should fall ~1.8 - 2 and give scientists an indication of purity versus...contaminants. The 260 and 230 nm ratios should fall ~2 - 2.2, and provide an indication of purity versus ...hydrochloride used during plasmid extraction. Figure 2: Use cloning grade DNA for direct cloning of your ... on these premises we can classify a plasmid preparation in 3 different ways: Transformation grade DNA...and flawless cloning it is important that the preparation is pure from other nucleic acids (genomic DNA...DNA for each reaction. As a consequence, the preparations for this technique must be larger scale, to ...

  11. The Materials Science of Optogenetics Experiments

    Type
    Blog Post
    ...different steps and materials required for each [1, 2]. This protocol [3] provides, in exquisite detail,...stimulation         In the video above, Channelrhodopsin 2 (Chr2) has been expressed in the motor cortex of a...lasers are relatively cheap and will run you about $2-3,000/laser. However, an important caveat is that ... the light-gated cation channel, Channelrhodopsin 2 (ChR2), exists in several states: closed/light receptive...PMID: 21628464; PubMed Central PMCID: PMC3138296. 2. Sparta DR, et al. Construction of implantable optical...board, Jacqueline N Crawley  [et al]. 2012;Chapter 2:Unit2 16. doi: 10.1002/0471142301.ns0216s58. PubMed...region. Thankfully, due to their prominence, the preparation of the fiber optics is probably the least difficult...

  12. Viral Vectors 101: AAV Serotypes and Tissue Tropism

    Type
    Blog Post
    ...N-linked sialic acids1 AAV receptor (AAVR)2   AAV2 Heparan sulfate proteoglycan (HSPG)3 AAV receptor...-linked sialic acids1 Heparan sulfate proteoglycan (HSPG)3 AAV receptor (AAVR)2 Epidermal growth factor...receptor (AAVR; not strictly required)2 Human fibroblast growth factor receptor 1 (FGFR1)3 αVβ5 and α5β1...AAV5 2,3-N-linked sialic acid4 AAV receptor (AAVR)2 Platelet-derived growth factor receptor (PDGFR... AAV8 Laminin receptor (LR)3 AAV receptor (AAVR)2 Carboxypeptidase D (CPD), also called AAVR25   ... Terminal N-linked galactose3 AAV receptor (AAVR)2 Laminin receptor (LR)3 Putative integrin3 ... AAV2, so they are formally called rAAV2/1, rAAV2/2, and so on, although we often use abbreviated nomenclature...

  13. Viral Vectors 101: Producing Your rAAV

    Type
    Blog Post
    ...digital PCR. Human Gene Therapy Methods, 25(2), Article 2. https://doi.org/10.1089/hgtb.2013.131 Lock,...Snyder, R. O. (2010). Characterization of a recombinant adeno-associated virus type 2 Reference Standard ... virus Once you have transfected your cells, wait 2–5 days and then collect the supernatant and/or producer...quantification of adeno-associated virus serotype 2-derived inverted terminal repeat sequences. Human ..., R., & Kleinschmidt, J. (1999). Titration of AAV-2 particles via a novel capsid ELISA: Packaging of genomes... genomes can limit production of recombinant AAV-2. Gene Therapy, 6(7), Article 7. https://doi.org/10.1038...been developed to determine the titers of rAAV preparations, such as dot blot, enzyme-linked immunosorbent...

  14. Plasmids 101: Secondary Nanobody Toolbox

    Type
    Blog Post
    ... larger label displacement Require separate incubation of 1° and 2° Must use different species and/or...antibody has an antigen binding site for a total of 2 binding sites per antibody. C) IgG’s are big enough...enough that multiple labels (green circles in figure 2 below) can be attached to each antibody. But being...Greater resolution due to lower label displacement 2° incubation can be skipped by pre-binding to 1° ...a fluorescent tag and the target antigen to about 2 nm, which can lead to higher resolution for super-...subclass.   Express the Nanobody In Bacteria: Table 2 also has a link to the plasmid you’ll need to express... Find plasmids from this publication at Addgene. 2. Bates, M., Böhning, M.I., Chug, H., Görlich, D., ...

  15. Five Popular Model Organisms, Part 2

    Type
    Blog Post
    ... H. J. (2009). Rats! Disease Models & Mechanisms, 2(5–6), 206–210. https://doi.org/10.1242/dmm.002733 ...is as a biomedical model organism.  Arabidopsis thaliana Arabidopsis thaliana is one of, if not the most...scientific history, highly characterized genome, and widespread utilization, Arabidopsis has made significant...community.  The Arabidopsis genome project marked the beginning of widespread Arabidopsis genetics research...world worked to characterize each gene’s function. Some key genetics research with Arabidopsis include understanding...feature genetic, physiological, or developmental characteristics that make them ideal for studying in a laboratory...genome has an extensive research history and characterization. Many proteins and processes naturally found...

  16. Adenoviral Vector Production and Troubleshooting

    Type
    Blog Post
    ...repeated multiple times (2-4 rounds) at increasing scale over the course of 1-2 weeks. Each round of amplification...creating adenoviral vector constructs. It consists of 2 plasmids: a shuttle vector (in which the transgene...Addgene’s Adenoviral Guide webpage, and references 1, 2.*Pro-Tip*: once the correct recombinant pAdV plasmid...sequenced to confirm its integrity. Initial production (2-3 weeks) - Here you’ll produce the primary recombinant...media will turn yellow. Do NOT change the media (add 2-3 mL of fresh media once a week), and do NOT harvest...it will result in very low titer. Amplification (1-2 weeks) - The rAdV-S is used to infect more HEK293 ...in a 10-100-fold increase in virus. Purification (2 days) - Purification is required if rAdV is to be ...

  17. Deep Mutational Scanning with One Pot Saturation Mutagenesis

    Type
    Blog Post
    ... that results in a polymerase error rate of up to 2% per nucleotide position. The “sloppy” PCR reaction...modifications: 1) increased concentration of Taq polymerase, 2) increased PCR extension time, 3) increased concentration...to synthesize a complementary mutant strand (step 2 and 3). Then the uracil-labeled template strand is...restrict site matter? Most of the time no, but there are 2 instances where it does. 1) When the backbone vector...ExoIII. See the below figure for an example like this. 2) When your gene of interest has a BbvCI site(s). If...saturation mutagenesis method are 1) ease of use, 2) time savings, and 3) ability to perform single site...PMID: 23766324. PubMed Central PMCID: PMC4409680. 2. Mccullum, E. O., Williams, B. A., Zhang, J., & Chaput...

  18. Cpf1: A New Tool for CRISPR Genome Editing

    Type
    Blog Post
    ...Cpf1 is a single RNA-guided endonuclease of a class 2 CRISPR-Cas system. Figure from Zetsche et al., 2015...2015. How was Cpf1 found and tested? Class 2 CRISPR systems, including the type II Cas9-based system...employed by class 1 systems. A putative new class 2 nuclease, Cpf1 (CRISPR from Prevotella and Francisella... the 3’ end of the target sequence. Figure 2: Effects of (A) double and (B) single base pair gRNA-target...Cpf1 is a single RNA-guided endonuclease of a class 2 CRISPR-Cas system." Cell (2015). PubMed PMID: 26422227... Find plasmids from this publication at Addgene. 2. Zetsche, Bernd, et al. "Multiplex Gene Editing by...al. added to the CRISPR toolbox with their characterization of two Cpf1 orthologs that display cleavage...

  19. AAV Titers: Where Do They Come From And What Do They Mean?

    Type
    Blog Post
    ...PubMed Central PMCID: PMC3640491. 2. Grimm, D., et al. "Titration of AAV-2 particles via a novel capsid ELISA...Lock, Martin, et al. "Characterization of a recombinant adeno-associated virus type 2 Reference Standard ...report the physical titer of a viral solution (Figure 2). Because physical titers are used for dosing purposes...mean and how they can be compared.   Figure 2: Label that comes on Addgene’s AAV aliquots. Our titer...of genomes can limit production of recombinant AAV-2." Gene therapy 6.7 (1999). PubMed PMID: 10455443. ...methods measure different characteristics of the solution. These characteristics give us information on ...vector preparations. Addgene has used this method to generally validate our viral vector preparation protocol...

  20. Multiplexed Capture of Promoter-enhancer 3D Chromatin Structures Using CRISPR

    Type
    Blog Post
    ...) FLAG-biotin-tagged dCas9, 2) BirA, and 3) one or more gRNAs Two separate lentiviruses delivering 1) ...Originally published Dec 7, 2017 and updated Jul 2, 2020. Promoters may be the star of gene regulation...sequence targeted in the experiment. Figure 2: Comparing CAPTURE and CAPTURE 2.0.     Original...) a BioTAP-tagged dCas9 and an eGFP tag, and 2) gRNAs Number of loci targeted One Many Sample type...identified and the level of on-target enrichment. 2. Determining the spatial and hierarchical organization...Cytogenetics, 11. https://doi.org/10.1186/s13039-018-0368-2  Liu X, Zhang Y, Chen Y, Li M, Zhou F, Li K, Cao H...chromatin interactions at a promoter in the well-characterized beta globin locus. The two methods had similar...
Showing: 61 - 80 of 490 results