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We narrowed to 815 results for: ache

Showing: 121 - 140 of 815 results

  1. Hot Plasmids April 2018 - Protein Degradation, Nanoscopy, FIRE-Cas9, and Yeast Expression Tools

    Type
    Blog Post
    Published
    April 23, 2018, 2 p.m.
    ...including difficult-to-manipulate primary cells, achieving knockdown within 1-2 hr of reagent delivery. TRIM-Away... sometimes incorporating chemically inducible approaches to provide temporal control, this new system,...Epigenome editing by Cas9 (FIRE–Cas9), combines these approaches to enable rapid and reversible recruitment of...target multiple loci at once. In addition, the approach allows minute-by-minute analysis of the kinetics...transcription factor as well as reporters specific to each transcription factor. They then induced cultures...induction via estradiol and progesterone do not affect each other, these systems can be used together. Aranda-Diaz... gene expression regimes that cannot be easily achieved with classical gene overexpression and knockout...

  2. Channelrhodopsins with improved light sensitivity for minimally-invasive optogenetics

    Type
    Blog Post
    Published
    Nov. 3, 2020, 2:15 p.m.
    ... Arnold’s lab and Viviana Gradinaru’s lab used machine learning to design new channelrhodopsins that are...channelrhodopsin that’s successful in vivo. Using machine-learning to overcome channelrhodopsin design challenges...challenges, the Arnold and Gradinaru Labs developed a machine-learning model to help them construct “designer...packaged in PHP.eB, ChRgers and ChR2(H134R) both reach the brain, but individual neurons do not express... mice. A fiber-optic cable allows blue light to reach the brain. Created with BioRender.com. In the...implanted into the mice’s brains so that light could reach the VTA. Mice were then allowed to “self-stimulate...Robinson JE, Mackey ED, Gradinaru V, Arnold FH (2019) Machine learning-guided channelrhodopsin engineering enables...

  3. Choosing the B(right)est Fluorescent Protein: Aggregation Tendency

    Type
    Blog Post
    Published
    June 15, 2017, 2:30 p.m.
    ...thanks to our guest bloggers Joachim Goedhart and Marieke Mastop! Joachim Goedhart is an assistant professor...This post was contributed by guest bloggers Joachim Goedhart and Marieke Mastop from the Section of Molecular...two posts in this series described a practical approach to selecting a bright fluorescent protein and ...homodimerization is characterized by an affinity of 110 µM (Zacharias et al, 2002), meaning that at this concentration...cells. To understand how well fluorescent proteins approach this ideal situation, cellular assays have been...fluorescent probes. You can follow him on twitter: @joachimgoedhart.   Marieke Mastop is a PhD candidate at the...

  4. A Look at Addgene's QC Process

    Type
    Blog Post
    Published
    May 6, 2025, 1:15 p.m.
    ...intended. Sharing unverified plasmids can lead to far reaching consequences, from wasting time and money on optimizing...Depositing scientists provide different types of data for each of their plasmids, from descriptions of key components...matter the level of information that is provided, each deposit goes through the same QC process to ensure...deposited sequences and plasmid information.  For each plasmid deposited at Addgene, QC scientists confirm...selectable markers, and fusion proteins listed for each plasmid  If the plasmids pass all inspection points...mutated; elements that are difficult to sequence; approaches to confirm plasmid sequences without a reference...confirm their presence with confidence. If this approach does not resolve the identified differences, the...

  5. Antibodies 101: Flow Cytometry

    Type
    Blog Post
    Published
    July 20, 2021, 1:15 p.m.
    ...process termed hydrodynamic focusing. This allows each cell to pass through the path of the laser in single-file...fluorescent light sources. Visible light illuminates each cell as it passes through and scatters in different...to a voltage pulse, called an event, and assigns each event to a channel number based on intensity. Higher...experiment uses 2 or 3 different fluorescent colors each measuring a different target. Flow cytometers however...stain the intracellular target. When using this approach, avoid fixation-sensitive fluorochromes such as...charged as they pass through a metal deflection plate. Each droplet contains a single cell that is assessed ...collection vessels and can sort single cells into each well allowing users to easily isolate and subsequently...

  6. Adapting Toehold Switches to Detect Zika Virus

    Type
    Blog Post
    Published
    Aug. 30, 2016, 2:30 p.m.
    ...need for low-cost, field-based diagnostics, our approach could make an impact. Our motivation was underscored.... We also were careful to set up clear tasks for each member of the team to accomplish so that many elements...expertise of the group and yet never hesitated to reach out to the greater research community when we needed...involved in the effort. This inclusive and open approach by all involved definitely contributed to our ...). Using toehold switch-based RNA sensors, our approach uses portions of the pathogen’s genome as unique...portable quantification of diagnostic results.  Our approach now detects samples at clinically relevant concentrations...longer-term aim is to advance our programmable approach to other critical needs and to further develop...

  7. PhD Applications After COVID

    Type
    Blog Post
    Published
    Jan. 5, 2023, 2:15 p.m.
    ...application cycle becomes much more complex. “How do I reach out to potential mentors?” “Where do I start with...research that aligns with your interests and goals.  Reaching Out to Potential PIs Even if you’re applying to...application process. Many institutions recommend that you reach out to potential advisors before submitting your...your application that you intend to work with, so reach out to three in case one isn’t able to respond to...your email. During my PhD application cycle, I reached out to around twelve professors and heard back ...you determine if the program is the right fit. Reaching out to PIs also means doing plenty of mini-interviews...applications can get to know faculty, students, and each other. Before COVID-19, financial and logistical...

  8. Golden Gate Assembly Upgrades: More Fragments, Faster Assembly, and Higher Fidelity

    Type
    Blog Post
    Published
    Oct. 11, 2018, 12:30 p.m.
    ...hierarchical approaches using different Type IIS restriction enzyme specificities at each step. Factors...Our work demonstrates that it is now possible to achieve 20+ fragment assemblies with both robust efficiency...Additional confirmation of accurate assemblies was achieved by the sequencing of plasmids isolated from blue...illustrates how the volume of the 1 ml outgrowth spread on each transformation plate can be manipulated to result...fragment lacI/lacZ cassette assembly was easily achievable with high levels of transformants and low backgrounds...researchers continue to test the limits of DNA assembly approaches with increasingly complex experimental conditions...

  9. Fluorescent Proteins 101: Fluorescent Protein Timers

    Type
    Blog Post
    Published
    May 4, 2017, 2:30 p.m.
    ...Snapp’s blog post shows. To avoid these issues, Subach et al. and Tsuboi et al. developed monomeric FP...mK-GO from Tsuboi et al. and the FT series from Subach et al.) were derived from previously developed ... while attempting to enhance mKO in other ways, Subach et al made a concerted effort to develop their ...see Table 1 for emission and absorption spectra). Subach et al’s directed mutagenesis also produced the ...Monomeric FP Timers and Associated Plasmids from Subach et al. Protein Excitation (nm) Emission (nm)...following its subcellular localization overtime (Subach et al.) Monitor protein expression dynamics in ... (2001): 29621-29624. PubMed PMID: 11408473. 4. Subach, Fedor V., et al. "Monomeric fluorescent timers...

  10. Google Forums Round Up: First Impressions of NgAgo

    Type
    Blog Post
    Published
    Aug. 4, 2016, 2:30 p.m.
    ... that many researchers were having difficulty achieving indel formation (gene disruption) with this new...used by respondents (transfection method, amount of each component per transfection etc.). Take Pooran's ...most of the respondents have been struggling to achieve genome editing with NgAgo, this does not mean the...successful indel generation and epitope knockin was achieved by a few respondents suggesting that NgAgo can...most of the researchers surveyed here could not achieve indels with the NgAgo system indicates that it ...which might be rapidly degraded by the cellular machinery, leaving a small amount of active NgAgo/ssDNA ... cell. As proposed in the original article, to achieve greater efficacy, researchers will have to carry...

  11. CRISPR Antimicrobials

    Type
    Blog Post
    Published
    May 3, 2016, 2:30 p.m.
    ...resistance is upon us, and the world is unprepared. Each year in the United States, two million people will...to combat antibiotic resistance using multiple approaches. Major restaurant chains including Chipotle and...bacteria have poor non-homologous end joining machinery, so a CRISPR-induced double-stranded break (DSB...is efficient delivery, as the antimicrobial must reach essentially every cell or else small pools of antibiotic...into the bacterial genome, and it carries CRISPR machinery targeting 1) the given resistance gene and 2) ...with the caveat that phages are very diverse, and each phage has a narrow bacterial host range. To further...Ando et al. recently engineered synthetic phages, each based on the well-studied T7 phage, to target various...

  12. CRISPR 101: Any Base Transversion Editors

    Type
    Blog Post
    Published
    April 1, 2025, 1:15 p.m.
    ...incorporate A bases across from abasic sites. This approach resulted in a maximum A → T product purity of ...AYBEv3!). Although AYBE, AXBE, and ACBE do not approach the high product purity of CBEs and ABEs, they...). Several different teams succeeded with this approach around the same time (He et al., 2024; Tong et...al., 2024; Ye et al., 2024; Yi et al., 2024). In each case, a UNG enzyme was modified to recognize T and...editing window than existing CGBEs. This distinct approach means researchers can now select the best tool...although the efficiency, purity, and indel rate of each tool vary considerably. And there’s so much more...on in the base editing field! From CRISPR-free approaches to mitochondrial base editing to base editors...

  13. Twenty Years of Sharing: Addgene's Viral Vector Service

    Type
    Blog Post
    Published
    July 30, 2024, 1:15 p.m.
    ...easy to ship as plasmids, they still have a global reach. From 2016–2023, we shared 93,491 viral vector preps...Cumulative AAV preps available in the Addgene repository each year from 2016–2023. B) AAV preps distributed per...viral vectors. This process builds on our former approach and involves reading primary literature, soliciting... viral vectors service puzzle: the usefulness of each viral prep in practice. The number of requests shows...redesigned Data Hub in 2022. It’s influenced the way we approach our viral vector educational resources, from eBooks...Cepko, C., Guerin, K. I., Rego, M. A., Ersing, I., Bachle, S. M., Kamens, J., & Fan, M. (2019). Adeno-Associated... Rachel Leeson...

  14. CRISPR Activation: A Practical Guide

    Type
    Blog Post
    Published
    Aug. 18, 2016, 2:30 p.m.
    ...specific UV wavelengths or chemicals (Polstein and Gersbach, 2015; Zetsche et al., 2015). These types of CRISPR...native conditions, the less activation you can achieve using a CRISPRa. Your gene of interest might already...your system is working as expected. Finally, for each gene you want to activate, you should also be ready...Dow D, Buser-Doepner C, Bertotti A, Trusolino L, Stronach EA, Saez-Rodriguez J, Yusa K, Garnett MJ (2019...engineering parameters via a library-on-library approach. Nat Methods 12:823–826 . https://doi.org/10.1038...JL, Davidsohn N, Housden BE, Perrimon N, Weiss R, Aach J, Collins JJ, Church GM (2015) Highly efficient...doi.org/10.1016/j.cels.2018.04.011 Polstein LR, Gersbach CA (2015) A light-inducible CRISPR-Cas9 system...

  15. Multicolor Animals: Using Fluorescent Proteins to Understand Single Cell Behavior

    Type
    Blog Post
    Published
    March 5, 2019, 1:08 p.m.
    ...colors to label an organ or entire organism. Because each cell can have multiple copies of the transgene that... colors when a combination of XFP are expressed. Each cell remains the same color for its entire lifetime... as many as 16 copies of the injected transgene, each of which could recombine to activate the expression...expression of a different fluorescent protein. Therefore, each cell could express a unique combination of all of...construct, dBrainbow includes small epitopes fused to each XFP. Secondary antibodies that target these epitopes...onto a tamoxifen-inducible ubiquitin background, each cell in the mouse that expresses Cre undergoes exactly... exactly one recombination event, leading to each copy of the transgene expressing either YFP or CFP, ...

  16. Site Directed Mutagenesis by PCR

    Type
    Blog Post
    Published
    Aug. 2, 2016, 2:30 p.m.
    ...substitutions (or deletions) in a tailored manner. The approach can be used in conventional cloning (to introduce...reverse primers are designed to be complementary, but each primer may extend beyond the complementary region...larger band (not shown). As a footnote; a similar approach can be used to identify Cas9/CRISPR-induced genome...template for the mutagenesis reaction. Alternative approaches to site directed mutagenesis Point-mutagenesis...PCR-introduced mutations can make alternative approaches more favorable if you want to introduce a point... introduce a frame-shift mutation. This can be achieved using a restriction enzyme that recognizes a unique...directed mutagenesis is one of the most popular approaches for introducing minor modifications to a plasmid...

  17. Evolution of Brainbow: Using Cre-lox for Multicolor Labeling of Neurons

    Type
    Blog Post
    Published
    April 24, 2015, 2:39 p.m.
    ...antibodies specific to each fluorophore. Third, Brainbow-1 and Brainbow-2 each contained a “default” state...construct contains two tandem invertible segments, each with two fluorescent protein coding sequences in...then successfully generated custom antibodies to each of these proteins and confirmed that there was no...opening the door for immunostaining analysis. To achieve even cell labeling, they generated farnesylated...-EF1a-BbTagBY. These constructs contain two XFPs each due to size limitations associated with AAV; co-... Nov 1;450(7166):56-62. PubMed. A technicolour approach to the connectome. Lichtman JW, Livet J, Sanes...

  18. Six Spooky Science Stories and Halloween at Addgene

    Type
    Blog Post
    Published
    Oct. 31, 2019, 1:06 p.m.
    ...gills. It attaches there until they mature and changes sex from male to female. Then, it detaches from the...their foraging trail. They climb upwards until they reach a point where the temperature and humidity favors...Vampirovibrio chlorellavorus is a predatory bacteria that attaches to the cell wall of green algae (Soo et al., 2015...eyestalks. Once ingested by the snail, the larvae reach the eyestalk and transform them into a pulsating... grasshopper harbouring hairworm: a proteomics approach." Proceedings of the Royal Society B: Biological...

  19. Tag Your Favorite Yeast Genes with Ease

    Type
    Blog Post
    Published
    Nov. 19, 2013, 2:37 p.m.
    ... MS, McKenzie A 3rd, Demarini DJ, Shah NG, Wach A, Brachat A, Philippsen P, Pringle JR. Yeast. 1998 Jul... in epitope tags? Others may be interested in attaching epitope tags to their genes of interest, allowing...complete collection of yeast tagging modules with each possible combination of protease site (TEV or PreScission...and selection marker (KanMX, HphMX, or His3MX).  Each PCR product from this collection will yield an insert...Longtine MS, Shah NG, McKenzie A 3rd, Steever AB, Wach A, Philippsen P, Pringle JR. Yeast. 1998 Jul;14(...

  20. Mapping the 4D nucleome with CRISPR/Cas9

    Type
    Blog Post
    Published
    Aug. 11, 2015, 2:30 p.m.
    ...could then be used to examine both inter- and intrachromosomal dynamics, and how they change in response ... to SpCas9 and its orthologs NmCas9 and St1Cas9. Each ortholog was fused to a different fluorescent protein... to differences in the PAM sequences required by each ortholog, a gRNA designed for one dCas9 should be...turned their attention to mapping pairs of intrachromosomal loci. The technique successfully resolved ... this work represents the first mapping of intrachromosomal loci, a major benchmark in characterizing ...under lower illumination settings, lowering photobleaching and phototoxicity concerns and extending the...
Showing: 121 - 140 of 815 results