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We narrowed to 520 results for: abo.2

Showing: 21 - 40 of 520 results

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

    Type
    Blog Post
    Published
    Jan. 11, 2017, 3:30 p.m.
    ...that increase cancelled. Rounding out the remainder, 2% of postdocs who weren’t going to get a salary increase...with their plans, and 17 have cancelled their plans. 2 of those raising - the University of Illinois and ...Previous Article: Changing Labor Laws Bring Increased Postdoc Wages Read about the Advocating for Science... of $47,476 per year, under updates to the Fair Labor Standards Act (FLSA). This was due to the threshold...postdocs - and in some cases, faculty - have found out about what their institution is doing from Future of Research...Science Symposium and Workshop Learn about a Variety of Science Careers Resources on Addgene.org Find Career...

  2. Giving gRNAs a Facelift - Synthetic and Beyond

    Type
    Blog Post
    Published
    Oct. 20, 2022, 1:15 p.m.
    ...same site, is 2’-fluoro (2’-F), which substitutes a fluorine for the hydroxyl group at the 2’ position. ...most common of these is 2’-O-methylation, where a methyl group is added to the 2’ hydroxyl group of the...them together, for example phosphorothioates with 2’-O-methyl modifications, proved more stability than...generated by introduction of a single photocleavable 2-nitrobenzyl linker in the 20-bp targeting region of...“I hope it isn’t degraded” when scientists talk about their precious RNA samples. RNA is inherently less...efficacy as well as the unnecessary stress of worrying about the status of your RNA. gRNAs modifications are ...Some of the RNA stabilizing modifications listed above were developed before CRISPR-Cas9, for use in applications...

  3. Great Results Start with Great Standard Curves

    Type
    Blog Post
    Published
    Aug. 15, 2024, 1:15 p.m.
    ... Figure 1 depicts a 2-fold dilution series. Each step of the series dilutes by 1:2 for a series that ranges...series ranging from 1:2 to 1:32. Each individual step of the series is consistent at 1:2. When preparing a.... An example of this is shown in Figure 2.   Figure 2: The concentration of a serially diluted... ranges from 1:2–1:32. The specific dilution series used will depend on the expected concentration of ...accuracy of the curve, avoid pipetting small volumes (< 2 µL) or volumes too large for a standard micropipette...by replicate values provides useful information about the accuracy of the curve, which we will discuss...many scientific applications users aim for an R2 above 0.95.    Additional considerations When running ...

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

    Type
    Blog Post
    Published
    Dec. 15, 2016, 3:57 p.m.
    ...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:...

  5. Typing CRISPR Systems

    Type
    Blog Post
    Published
    March 18, 2025, 1:15 p.m.
    ...   Class 1 versus class 2 There are two CRISPR classes: Class 1 and Class 2. Class 1 CRISPR systems use...systems to carry out various functions. Class 2 Class 2 CRISPR systems rely on a single protein Cas effector...explore Class 2 CRISPR in more depth, the Jennifer Doudna lab maintains an encyclopedia of Class 2 CRISPR systems...systems: a burst of class 2 and derived variants. Nature Reviews Microbiology, 18(2), 67–83. https://doi.org...complex for antiviral defense) complexes, while Class 2 use single protein Cas effectors, like Cas9. The Cascade...1 remains largely underutilized compared to Class 2. Class 1 Despite their incredible abundance, Class...components present in the Cascade complex. Figure 2: Schematic of a Cascade complex (blue subunits) with...

  6. Plasmids 101: Knockout/Knock-In Plasmids

    Type
    Blog Post
    Published
    Dec. 1, 2016, 3:30 p.m.
    ...the addition of Cre recombinase (Figure 2). Figure 2: A knockout targeting vector designed to... through an experiment designed to knock out exon 2 of a given gene.   Figure 1: A knockout targeting...For recombination to occur in a cell, a minimum of 2 kb of sequence homology is required, but 6 to 14 kb...construct to your target cells. After recombination, exon 2 of the targeted gene will be removed from the chromosome...the GFP/NeoR cassette is inserted in place of exon 2. Thus, the targeted gene is disrupted (knocked out...recombination event will remove NeoR only and leave exon 2 floxed, as seen in line 4 of Figure 4. Since the loxP...inserted GFP is expressed (knocked in). As seen in the above example, you can remove a floxed resistance gene...

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

    Type
    Blog Post
    Published
    March 7, 2017, 3:30 p.m.
    ...designed (listed above) worked well and I got clear bands using the conditions above. 2. Design your primers...expression, but I didn’t. 2. In the original publication they see GFP expression in about 20% of their Cas9-expressing...transductions and use a range of reasonable MOIs. 2. Create a monoclonal Cas9-expression cell line. Especially...doing a screen. I made a few monoclonal lines (Figure 2) and I was surprised at how variable Cas9-expression...across the different cells (Figure 3).   Figure 2: Cas9 expression in monoclonal cell lines generated... expression Overview: By Western blotting (Figure 2, pool), I knew the cells expressed Cas9, but I didn... appear green despite having active Cas9 (see tip 2). Pro-Tips 1. To quantify Cas9 activity using this...

  8. Plasmids 101: Secondary Nanobody Toolbox

    Type
    Blog Post
    Published
    Feb. 27, 2018, 2:04 p.m.
    ...between a fluorescent tag and the target antigen to about 2 nm, which can lead to higher resolution for super-resolution...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...displacement Require separate incubation of 1° and 2° Must use different species and/or different IgG ...Greater resolution due to lower label displacement 2° incubation can be skipped by pre-binding to 1° ...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., ...

  9. Rosella: A Fluorescent pH-Biosensor for Studying Autophagy

    Type
    Blog Post
    Published
    April 13, 2017, 2:30 p.m.
    ...Table 2 for more details on the effect of pH on the fluorescent properties of Rosella.   Table 2: Effect...in yeast. Autophagy,4(2), 205-213. doi:10.4161/auto.5331. PubMed PMID: 18094608 2. Sargsyan, A., Cai, J...environment like the vacuole or lysosome. See Table 2 for a summary of what color fluorescence Rosella emits...fluorescence accumulates in the vacuole. See Figure 2 for an example of what fluorescence looks like for...information about the contents of the autophagosome nor does it provide information about what is being...higher pH of the vacuole. Read on to learn more about prior methods for studying autophagy and how Rosella... (as opposed to giving more fine-grained detail about what proteins or organelles are being degraded) ...

  10. Hot Plasmids - October 2022

    Type
    Blog Post
    Published
    Oct. 6, 2022, 1:15 p.m.
    ...GEVIs (called ASAP 1 and 2), they found an indicator that is optimized for 2-photon imaging of voltages...resultant JEDI-2P sensors overcome many challenges in 2-photon in-vivo imaging of voltage fluctuations. Image...suitable for in vivo optogenetic use.   Fig. 2: A) HcKCR1 is highly selective for K+ over other Na...slice of HcKCR1-EYFP and tdTomato expressed layer 2/3 neurons in mouse. C) Action potential frequencies...Brian O'Neill  The François St-Pierre lab and collaborators have successfully developed an improved voltage...

  11. The Materials Science of Optogenetics Experiments

    Type
    Blog Post
    Published
    Sept. 17, 2015, 2:30 p.m.
    ...Laser stimulation         In the video above, Channelrhodopsin 2 (Chr2) has been expressed in the motor...These lasers are relatively cheap and will run you about $2-3,000/laser. However, an important caveat is that...different steps and materials required for each [1, 2]. This protocol [3] provides, in exquisite detail,... 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...distance so ideal implant placement should be about 0.4-0.5mm above the site of your stimulation. The scatter...

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

    Type
    Blog Post
    Published
    June 16, 2020, 1:25 p.m.
    ... 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 ... cloning Read our Plasmids 101 blog posts Learn about the differences between transformation, transduction...

  13. Viral Vectors 101: Producing Your rAAV

    Type
    Blog Post
    Published
    July 16, 2024, 1:15 p.m.
    ...digital PCR. Human Gene Therapy Methods, 25(2), Article 2. https://doi.org/10.1089/hgtb.2013.131 Lock,... 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...Characterization of a recombinant adeno-associated virus type 2 Reference Standard Material. Human Gene Therapy, 21...dehydration (Howard & Harvey, 2017). You can learn more about common AAV applications by reading our Viral Vectors...

  14. Deep Mutational Scanning with One Pot Saturation Mutagenesis

    Type
    Blog Post
    Published
    Feb. 22, 2017, 3:30 p.m.
    ...image above). A second universal primer is used to synthesize a complementary mutant strand (step 2 and ... 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...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...

  15. CRISPR Between the Genes: How to Experiment with Enhancers and Epigenomics

    Type
    Blog Post
    Published
    Aug. 9, 2016, 2:30 p.m.
    ...for the 2+ million DHS sites. See below for further details. DNA is a 3D Fractal Figure 2: DNA compaction...: 1. Find Enhancers that have NANOG binding sites 2. Design a CRISPR screen to target and disrupt the ...NANOG enhancers 3. CRISPR out the enhancers from step 2. See which ESCs die or differentiate Step 3 reveals...expression for NANOG enhancers, we’re ready for step 2: design a CRISPR screen to disrupt these enhancers...to CRISPR out the enhancers we identified in step 2. The result is a heterogenous population of stem cells...republished here with permission. Quilt is a collaborative database for genomics. In this article, Quilt...tri_methylation In the next section we’ll apply the above formula to a real-world experiment. We’ll start ...

  16. Five Popular Model Organisms, Part 2

    Type
    Blog Post
    Published
    Sept. 29, 2020, 1:15 p.m.
    ... H. J. (2009). Rats! Disease Models & Mechanisms, 2(5–6), 206–210. https://doi.org/10.1242/dmm.002733 ... more about model organisms in the first Five Popular Model Organisms blog post Read more about plasmids...characteristics that make them ideal for studying in a laboratory environment. We’ve covered five model organisms...have been specifically developed to thrive in laboratory settings. One of its most attractive features...minutes, they are perfect for molecular cloning and metabolic engineering.  The E. coli genome has an extensive...address many areas of microbial research like metabolism. Transport, biofilms, and much more. Because ...characterized genome, and is easily maintained in a laboratory environment. What sets Arabidopsis apart from...

  17. Cpf1: A New Tool for CRISPR Genome Editing

    Type
    Blog Post
    Published
    Oct. 14, 2015, 2:30 p.m.
    ...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...genome editing than Cas9 - read on to learn more about Cpf1 and check out our CRISPR guide for a refresher...

  18. SciComm with the Experts at Science in the News Part 2

    Type
    Blog Post
    Published
    April 27, 2017, 2:30 p.m.
    ...Introduction - Vini and Amy tell us a little about themselves. Chapter 2: What is Science in the News (SITN)? ...To learn more about the STIN's podcast, email [email protected]    For more info about starting your...evens where scientists can grab a beer and chat about their work at a local bar. It doesn't have to be...science communicators. In this chapter we learn about some of their events - science seminars, science...the lab and working with SITN. She points to collaboration and delegating as keys achieving success. Chapter...your own SciComm organization and to learn about some of SITN's other initiatives, email [email protected]...

  19. Finding nucleic acids with SHERLOCK and DETECTR

    Type
    Blog Post
    Published
    April 16, 2020, 1 p.m.
    ... can be used to diagnose the presence of SARS-CoV-2 RNA, but inadequate access to reagents and equipment...named this system SHERLOCKv2 for SHERLOCK version 2. Here are some of the improvements: SHERLOCKv2 uses... a protocol for using SHERLOCK to detect SARS-CoV-2 RNA. The test is started using RNA purified from patient...recently adapted the DETECTR platform to detect SARS-CoV-2 using a lateral flow strip format. They published ...CY (2020) CRISPR–Cas12-based detection of SARS-CoV-2. Nature Biotechnology. https://doi.org/10.1038/s41587... and Functional Characterization of Diverse Class 2 CRISPR-Cas Systems. Molecular Cell 60:385–397 . https...Cpf1 Is a Single RNA-Guided Endonuclease of a Class 2 CRISPR-Cas System. Cell 163:759–771 . https://doi....

  20. Important Considerations When Using AAVs

    Type
    Blog Post
    Published
    June 13, 2017, 2:30 p.m.
    ...University of Manitoba due to their Biosafety Level-2 or 2+. Please Note: Despite the fact that AAVs themselves...long and stable expression (up to 9 months in mice [2]). Low Immune Response: Compared to other viral vectors... location. Location (1): What tissue? Location (2): Where in the tissue/which cell type? Location (3...may be sufficient to overcome the transport issues. 2. Failure to plan (for storage) Is planning to fail...Find Ready-to-Use Retrograde AAV Preps at Addgene 2) Comparison of genetically engineered receptors (i.e...Therapy 15.10 (2007): 1740-1743. PubMed PMID: 17882145. 2. Zincarelli, Carmela, et al. "Analysis of AAV serotypes... tropism and axonal transport of AAV serotypes 1, 2, 5, 6, 8 and 9 in the mouse brain." PloS one 8.9 (...
Showing: 21 - 40 of 520 results