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We narrowed to 297 results for: hal.2

Showing: 41 - 60 of 297 results

  2. 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 ...biomedical model organism.  Arabidopsis thaliana Arabidopsis thaliana is one of, if not the most, recognized...Arabidopsis genetics research development. Arabidopsis thaliana was fully sequenced by the year 2000, becoming...properties affecting adaptations.  Arabidopsis thaliana flowers. Image from BlueRidgeKitties. If you...functions in an ecological context with Arabidopsis thaliana. ELife, 4. https://doi.org/10.7554/elife.06100...

  3. Plasmids 101: Knockout/Knock-In Plasmids

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    Blog Post
    ...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...of interest is essential, a true knockout can be lethal, and you'd instead want to create a conditional...

  4. SARS-CoV-2/COVID-19 Detection Methods Based on CRISPR/Cas

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    Blog Post
    ... State University. Since its appearance, SARS-CoV-2 has spread to almost every part of the world manifesting...Rapid detection of 2019 novel coronavirus SARS-CoV-2 using a CRISPR based DETECTR assay Sensitivity:10-...primers that target the E and the N2 gene of SARS-CoV-2. If the Cas12a-sgRNA complex binds to the target sequences... method has been optimized to detect the SARS-CoV-2 virus directly using saliva without having to perform...and visual detection of novel coronavirus SARS-CoV-2 Sensitivity: 4.6-11 copies/microliter of RNA targets...single-molecule sensitive. In the case of HIV-1 and SARS-CoV-2 detection, AIOD-CRISPR without preamplification was...easy-to-deploy, protocol for Cas13-based detection of SARS-CoV-2 genetic material (CREST) Sensitivity: 10 copies of...

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

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    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...

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

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    Blog Post
    ...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...Western blotting against GFP expression, but I didn’t. 2. In the original publication they see GFP expression...and I got clear bands using the conditions above. 2. Design your primers so that the digestion site isn...

  7. Tips for Getting a Faculty Position

    Type
    Blog Post
    ... this problem? (2-3 sentences) What will be the focus of your first grant? (1-2 sentences) ...of scientific success is publications and grants. 2. Write succinct and easy to understand documents A...consideration. 3. Present an easy to follow seminar See item 2. Attendees will include grad students (that are solicited...people would agree is important to understand. (1-2 sentences). For example: Distinguishing correctly ...Talk." 4. The chalk talk, simple in principle The first question I usually get about chalk talks is, what... whether there is chalk. Sometimes. It can also be a slide deck, markers, etc. Chalk or markers force ... Developing your Chalk Talk These items should be part of the beginning of your chalk talk. These should...

  8. Adenoviral Vector Production and Troubleshooting

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    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 ...

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

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    Blog Post
    ...and Amy tell us a little about themselves. Chapter 2: What is Science in the News (SITN)? - SITN is a graduate...This is the second half of a two-part interview with Vini Mani and Amy Gilson from Science in the News...involved in science communication. In this second half of our conversation with Vini Mani and Amy Gilson...

  10. Great Results Start with Great Standard Curves

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    Blog Post
    ... 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...allergy testing lab, and the grad student down the hall from you have in common? All of them rely on standard...

  11. Rett Syndrome: A History of Research and Therapeutic Outlooks

    Type
    Blog Post
    ...//doi.org/10.1002/1096-8628(200022)97:2<147::aid-ajmg6>3.0.co;2-o Baker SA, Chen L, Wilkins AD, Yu P, ...Caused by a mutation in the methyl-CpG binding protein 2 (MECP2) gene, Rett Syndrome is a rare, progressive...X-linked MECP2, encoding methyl-CpG-binding protein 2. Nat Genet 23:185–188 . https://doi.org/10.1038/13810... (2000) Rett syndrome: Methyl-CpG-binding protein 2 mutations and phenotype-genotype correlations. Am ...cassette using tamoxifen-inducible Cre not only halted the neurological damage but completely reversed... Rett Syndrome, respectively. One of the main challenges facing Rett researchers is how to deliver the...adeno-associated viral (AAV) vector. The editase repaired half of the mutant MECP2 RNA in each of three neuronal...

  12. Harnessing TnpB for Genome Editing: A Compact and Versatile Tool for Your Research

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    Blog Post
    ...Oligo 2 to screen. A 667-bp product confirms successful insertion. Step 6: Confirmation Pick 2–3 positive...can use TEEP — a tool based on animal data. Step 2: Prepare Oligos Oligo 1: Add 'tcaa' to the 5′ end...your forward guide sequence (see Figure 1). Oligo 2: Add 'ggcc' to the 5′ end of your reverse complement...Primer 92F (5′-cattacgcaattggacgacaac-3′) and Oligo 2 (specific to your target) to screen colonies via PCR...), 84–89. https://doi.org/10.1038/s41587-019-0337-2  Molla, K. A., Sretenovic, S., Bansal, K. C., & Qi...approximately 1000–1400 amino acids — which poses a challenge for efficient delivery into cells, particularly...Cas12a. Applications of TnpB One of the major challenges in genome editing is the efficient delivery of...

  13. Hot Plasmids - October 2022

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    ...GEVIs (called ASAP 1 and 2), they found an indicator that is optimized for 2-photon imaging of voltages...The resultant JEDI-2P sensors overcome many challenges in 2-photon in-vivo imaging of voltage fluctuations...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...

  14. Typing CRISPR Systems

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    Blog Post
    ...   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...

  15. Optogenetics + CRISPR, Using Light to Control Genome Editing

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    Blog Post
    ...N-terminal (residues 2-713, N713) and C-terminal (residues 714-1368, C714) halves, rendering the Cas9 ...the light-inducible heterodimerizing cryptochrome 2 (CRY2) and calcium and integrin-binding protein 1 ...inactive, dead Cas9 protein (dCas9) fused to CIB1; and 2) the activator - the CRY2 photolyase homology region... fused to both the N- and C-termini of dCas9; and 2) CRY2FL-VP64, a fusion of full-length CRY2 and the...researchers saw an 11-fold increase in mRNA levels after 2 hr and a 400-fold increase after 30 hr. The system...nMagC714 instead of nMagHigh1-C714, generating paCas9-2. This change did not significantly alter the system...prior work, the Sato lab also showed that the paCas9-2 system could be spatially controlled and reversibly...

  16. 22 Hot Plasmid Technologies from 2014

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    Blog Post
    ...Currently available from the Kim lab are 2 presynaptic and 2 postsynaptic targeting mGRASP plasmids. Additionally... cells were pulsed with 2µg/ml of Doxycycline for 2 hours, and imaged on the Keyence BZ-9000 fluorescence...work! Robinson et al., Dev Cell 2010 Feb 16;18(2):324-31. Cheeseman et al., J Cell Sci 2013 May 1...promotes the formation of an isopeptide bond between 2 peptide tags, SpyTag and KTag. The group demonstrated... of three sets of cloning vectors (Level 0, 1, or 2) which can be utilized in three successive assembly...1 modules can be similarly assembled into a Level 2 vector, thus forming a functional genetic circuit....circuit. Flexibility has been built into the Level 2 vectors to allow for additional iterations of Level 1 assembly...

  17. Viral Vectors 101: Optogenetic Tools

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    Blog Post
    ...for controlling neuronal activity, channelrhodopsin-2 (ChR2), a blue light-activated cation channel, from...used in tandem with the opsin.      Figure 2: Diagram of excitatory (ChR2) activation by light ...) domains, phytochrome B (PhyB), and cryptochrome 2 (CRY2) (Zhang & Cui, 2015). When stimulated with light...comparative analysis of microbial opsins. Nature Methods, 9(2), 159–172. https://doi.org/10.1038/nmeth.1808 References...comparative analysis of microbial opsins. Nature Methods, 9(2), 159–172. https://doi.org/10.1038/nmeth.1808 Nagel...Hegemann, P., & Bamberg, E. (2003). Channelrhodopsin-2, a directly light-gated cation-selective membrane ...intracellular signaling pathways. Trends in Biotechnology, 33(2), 92–100. https://doi.org/10.1016/j.tibtech.2014.11.007...

  18. CRISPR Antimicrobials

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    Blog Post
    ...Citorik Plasmids here). In both cases, they used 1-2 CRISPR crRNAs targeting sequences found only in certain...resensitize antibiotic-resistant bacteria (Figure 2). The first phage introduced is lysogenic, integrating...machinery targeting 1) the given resistance gene and 2) a second lytic phage. Bacteria that receive this ...bacteria have previously flourished.   Figure 2: Two-phage strategies to resensitize bacteria to antibiotics...237163. Find plasmids from this paper at Addgene. 2. Bikard, David, et al. “Exploiting CRISPR-Cas nucleases...may provide a method for doing just that. While challenges remain in the delivery of these agents, CRISPR...CRISPR-induced double-stranded break (DSB) in the genome is lethal. If this DSB occurs on a plasmid, the plasmid will...

  19. Delivery Methods for Generating iPSCs

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    Blog Post
    ...transfection where 2 of 11 clones tested had plasmid integration. Another challenge is that multiple transfection...Recent progress and future challenges." Journal of Cellular Physiology 228, no. 2 (2012): 267-75. PubMed ...plasmid-based expression requires serial transfection of 1 or 2 plasmids that express the reprogramming factors of...understanding the mechanisms of reprogramming, and 2) studies with clinical endpoints. In the first scenario...PubMed PMID: 27764162. PubMed Central PMCID: 5072705. 2. Sommer, Cesar A., and Gustavo Mostoslavsky. "The ...PubMed PMID: 23546745. PubMed Central PMCID: 4176696. 2. Yusa, Kosuke, Roland Rad, Junji Takeda, and Allan...generation. Expert opinion on biological therapy, 10 2, 231-42. PubMed PMID: 20088717. PubMed Central PMCID...

  20. The Materials Science of Optogenetics Experiments

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    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...validation experiments we discussed last week are only half of the battle when using optogentics to answer a...
Showing: 41 - 60 of 297 results