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We narrowed to 53 results for: stem loop

Showing: 1 - 20 of 53 results

  1. Protocol - How to Streak a Plate

    Type
    Protocol
    ...bacterial colonies. Equipment Sterile toothpicks or wire loop Bunsen burner (or other small flame source) Incubator...bacterial stab or glycerol stock . Using a sterile loop, pipette tip or toothpick, touch the bacteria growing...of the glycerol stock. Pro-Tip If you use a wire loop you can sterilize it by passing it through a flame...flame, just be sure to allow enough time for the loop to cool before touching it to the bacteria. Gently... fresh, sterile toothpick, or freshly sterilized loop, drag through streak #1 and spread the bacteria ...third sterile pipette tip, toothpick, or sterilized loop, drag through streak #2 and spread the bacteria ...is recommended that you keep a standard labeling system for all your objects/solutions. Sterilize your ...

  2. A Needle in a Base-Stack: Cas9 Structural Biology

    Type
    Blog Post
    ...separating it into two strands: the stem loops and the guide region. The stem loops bind along the outside of the...doubles back on itself at the bottom of the image. Stem loops 1 and 2 extend horizontally across both lobes... guide RNA in this structure does not include stem loop 3, and guide RNA nucleotides 11-20 are not resolved... the protein, spanning both lobes. Stem loop 1, nestled between the REC and CTD domains, is required for... for Cas9 nuclease function. Stem loops 2 and 3 are not strictly necessary for in vitro cleavage, but ...strand are not resolved due to disorder. All three stem loops and all 20 guide bases of the RNA are shown. ...stabilizes the last base pair with an end-capping loop (Nishimasu et al., 2014). This cap is where the ...

  3. Hot Plasmids - September 2019 - CRISPR, Auxin Inducible Degradation, MerMAID Channelrhodopsins, and Genome Looping

    Type
    Blog Post
    ...synthetic looping, the Phillips-Cremins lab developed the light-activated-dynamic-looping (LADL) system. The...our chromatin is comprised of thousands of loops. These loops not only provide a 3-D structure for packing... endogenous looping on distal promoters and enhancers, or have attempted to engineer loops to create genomic...Communications, 2019. PubMed PMID: 31346176. Using LADL to loop with light Article contributed by Eric Perkins Mammalian...interactions that wouldn’t normally occur. The synthetic looping factors developed thus far for these latter experiments... are over 500kB apart, they were able to observe loop formation as well as increased Zfp462 expression...the dynamic nature of genomic looping due to it’s briefer time scale and its inducible and reversible nature...

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

    Type
    Blog Post
    ... eight surface-exposed loops of an mCherry nanobody. Only insertion into loops 1 and 6 resulted in light...opposite effects: insertion into loop 1 caused dark-induced binding while loop 6 insertion caused blue-light...AsLOV2 into loop 1 always generated OptoNBs with dark-induced binding, but insertion into loop 6 generated...mCherry #1 mCherry #2 GFP Loop 1 dark-induced dark-induced dark-induced Loop 6 blue-light induced dark-induced...binding of their target proteins. The surface-exposed loops of nanobodies seemed like a good place to put AsLOV2...membrane-localized mCherry or GFP protein. With this system, if the OptoNB can bind, it co-localizes with its...

  5. All in a Twist: dsRNA

    Type
    Blog Post
    ...prone to persistent R-loop formation, presumably due to their abundance of R-loops and repetitive sequence...the 3-stranded structure produced is known as an R-loop. During transcription, RNA is synthesized from ...intermediates in the process. These transcription-coupled R-loops are transient in nature and will dissociate when...these nucleic acid hybrids are always around. R-loops are particularly enriched in highly transcribed ...sequence. All of the R-loops described above play distinct roles in promoting cellular processes and functions...DNA strand. A ssDNA molecule exists within the R-loop that is now vulnerable to various forms of DNA damage...strike a balance between productive levels of R-loops and putting DNA at risk. Consequences of duplexed...

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

    Type
    Blog Post
    ...of gene regulation, but enhancers and chromatin looping play important supporting roles. Enhancers are ... regulate, but are brought in proximity by the looping of chromatin, the complex of DNA and proteins that...of 3C methods as taking a snapshot of chromatin looping with the photo “developed” by analyzing DNA sequencing...ends created are then ligated together to create loops of DNA. This is key to linking pieces of DNA that... populations. Key components of the CAPTURE 2.0 system To improve upon the original CAPTURE method, the...enzyme BirA Endogenous biotin ligases Expression system Stable cell line expressing 1) FLAG-biotin-tagged...

  7. Beyond the Blood Brain Barrier: PHP.B Enters the Central Nervous System Via the LY6A Receptor

    Type
    Blog Post
    ...Stanford. "Concise review: stem cell antigen‐1: expression, function, and enigma." Stem cells 25.6 (2007): 1339...AAV9 mutants that had a small randomized peptide loop inserted in the AAV9 capsid. This mutant library...Deverman and Wilson labs determined LY6A, also known as stem cell antigen-1 (SCA-1), is the receptor used by ...commonly used to enrich adult mouse hematopoietic stem cells. Its ligand(s) has not been identified. While...of LY6A, but a mutation in PHP.B’s unique capsid loop disrupted binding. Collectively these results suggest...barriers to gene therapy treatment of central nervous system (CNS) diseases is the blood brain barrier (BBB)...delivery to the central and peripheral nervous systems." Nature neuroscience 20.8 (2017): 1172. PubMed...

  8. AAV Molecular Tools

    Type
    Collection
    ...tet-off transactivator (tTA) with a positive feedback loop for amplified tTA expression. 1 Gradinaru 99121 ...tet-off transactivator (tTA) with a positive feedback loop for amplified tTA expression. 1 Gradinaru 117383...tetracycline (tet)-inducible expression systems . ID Name Expression System Activity Serotype PI 99118 pAAV-CAG-tTA... in the CRISPR-Cas9 system for targeted genome editing. ID Name Expression System Activity Serotype PI...can be used for viral TRAP). ID Name Expression System Activity Serotype PI 98747 pAAV-FLEX-EGFPL10a EF1a-driven...neuronal connectivity labeling. ID Name Expression System Activity Serotype PI 51509 AAV phSyn1(S)-FLEX-tdTomato-T2A-SypEGFP-WPRE...can be used for cell ablation. ID Name Expression System Activity Serotype PI 124364 pAAV-FLEX-DTR-GFP CBA-driven...

  9. Lighting Up Cell Signaling with Photoswitchable Kinases

    Type
    Blog Post
    ...Studying negative feedback loops of signaling pathways Negative feedback loops are a common feature of signal...potentially be created by inserting pdDronpa domains into loops of a protein of interest. If you plan to try this...kinases and use MEK1 as a model kinase. In this system, light is used to turn “on” and “off” MEK1 activity...cytoplasm. This all-optical psMEK1 + ERK KTR-mRuby2 system has several advantages over existing methods. First... or growth factors are used as a stimulus. This system also allows precise modulating of a particular ... kinases could possibly be used in other model systems, but they need to be expressed in a tissue or organism...

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

    Type
    Blog Post
    ...incorporating two MS2 RNA aptamers at the tetraloop and stem-loop 2; 3. The MS2-P65-HSF1 plasmid which expresses...The structural details revealed two exposed RNA loops, which Konermann recognized as an ideal spot to ...activation in mind. “We had not managed to create a system to allow us to reliably activate essentially any... to selectively activating them with the CRISPR system was a large one.  The question for her then was...it’s working. You can only do that screen with a system you are really confident about working for most...colleagues in Feng Zhang’s lab describe just such a system, which they call CRISPR/Cas9 Synergistic Activation... RefSeq database. The key to developing the new system, Konermann explained, was new knowledge about Cas9...

  11. Fluorescent Proteins 101: Aptamer Fluorophores

    Type
    Blog Post
    ...the stem in an RNA stem-loop structure (see Fig. 1a). When the aptamer binds its ligand, the stem loop...to target molecules. Using a technique called systematic evolution of ligands by exponential enrichment...fluorescence to create an improved fluorescent aptamer system. Knowing that GFP’s  structure stabilizes a 4-hydroxybenzylidene...8. Sassolas, Audrey, et al. “Optical detection systems using immobilized aptamers.” Biosens Bioelectron...

  12. Malate Dehydrogenase CUREs Community Collection

    Type
    Collection
    ... (specific mutation, "active site", "active site loop", or "subunit interface") ID Plasmid Description...Collaboration Results in Student Learning Gains and Positive STEM Attitudes in CUREs. Callahan KP et al. CBE Life ...

  13. CRISPR 101: Homology Directed Repair

    Type
    Blog Post
    ...DNA structure referred to as the displacement loop (D loop). The recombination intermediates can then ... Unlike in DSBR, following strand invasion and D loop formation, the newly synthesized portion of the ...based on your locus of interest and experimental system.  Synthesizing ssODNs longer than 200 bases can...efficiency varies across loci and experimental systems, ssODN templates generally provide the highest ...References: Genome engineering using the CRISPR-Cas9 system. Ran FA, et al. Nature Protocols Nov;8(11):2281...Genome Engineering Approaches with the CRISPR/Cas9 System. Li, K et al. PLoS ONE 9(8); (2014). PubMed. ...

  14. Hot Biosensors 2022: Year-End Roundup

    Type
    Blog Post
    ...region-to-IntrAcellular loop linkers” (MTRIA for short), where a fluorescent protein is inserted into one of the loop regions... a circularly-permuted GFP into an intracellular loop of various G-protein-coupled receptors. Image adapted...Toettcher and colleagues describe pYtags, a modular system for monitoring specific RTK activation in vivo ...

  16. Antibodies 101: Reading a Flow Plot

    Type
    Blog Post
    ... which can send you in an endless loop. Let’s break out of the loop with all you need to know on how to...Evaluation of in vitro chondrocytic differentiation: A stem cell research initiative at the King Abdulaziz University...

  17. Build Your CRISPR Vocabulary

    Type
    Blog Post
    ...antirepeat duplex. tracrRNAs also contain multiple stem loops that are recognized by the Cas9 protein from ...biology, too. CRISPR is an elegant bacterial immune system and an efficient gene editing tool… but boy does...bacterial chromosome encodes a tracrRNA (in some systems including Cas9), Cas proteins, and a CRISPR array...nontarget strand. Protein Cas: Different CRISPR systems involve different proteins, but many of them are... of DNA to be cut, which varies between CRISPR systems. Cas proteins won’t cut DNA that doesn’t have their...folds back on itself, simplifying the CRISPR-Cas9 system for laboratory applications. At Addgene, we sometimes...

  18. No Llamas Required - Synthetic Nanobodies Against Membrane Proteins

    Type
    Blog Post
    ...structures (concave, loop, and convex). Each of these 3 mRNA libraries have a concave, loop, or convex scaffold...Sybody Generation Toolbox plasmid kit: SB_concave, SB_loop, SB_convex. These scaffolds contain serines and...to 1012 members) in comparison to other display systems using phage and yeast (109-1010 members). This ...FX Cloning: A Versatile High-Throughput Cloning System for Characterization of Enzyme Variants. In: Methods...

  19. Degrading DNA with Cascade-Cas3

    Type
    Blog Post
    ...unwinding of dsDNA occur simultaneously, which creates a loop in the target strand that is cut occasionally as... Taylor, D. W. (2023). Structural snapshots of R-loop formation by a type I-C CRISPR Cascade. Molecular...editing. Class 1 CRISPR systems make up the majority of CRISPR-based immune systems in bacteria and archaea...Cascade-Cas3 plasmids in Addgene's collection! A “complex” system Cas3 is part of the Class 1 CRISPR family, which...genomic editing. Specifically, these types of CRISPR systems work using a multi-Cas complex called Cascade (...further broken down into different types, with Cas3 systems making up type I, the most abundant type in the...Cascade is an important component of the CRISPR Cas3 system. Most Cascade complexes are comprised of a combination...

  20. Using Phosphoserine to Study Protein Phosphorylation

    Type
    Blog Post
    ...requisite dual phosphorylation events on its activation loop, but can be made constitutively active through phosphomimetic...mentioned, phosphomimetic mutations on the activation loop do not fully activate MEK1. In fact, replacing just...on protein phosphorylation in an array of model systems coupled with advances in mass spectrometry suggest...kinase specificity in vitro. The establishment of a system in which purified, recombinant protein could be...researchers to create an orthogonal translation system (OTS) for the incorporation of phosphoserine directly... recombinant proteins (5). Optimization of this system defined three components required for translation...open reading frame of interest. Two expression systems have been verified for use by the Rinehart lab:...
Showing: 1 - 20 of 53 results