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We narrowed to 340 results for: abo.4

Showing: 1 - 20 of 340 results

  1. High Complexity Golden Gate Assembly with NEB’s DAD

    Type
    Blog Post
    Published
    Aug. 1, 2023, 1:15 p.m.
    ...cycling 4-base Bsal-HFv2 37-16 °C cycling 4-base BsmBI-v2 42-16 °C cycling 4-base ... 1 hour 3-base or 4-base T4 DNA ligase 25 °C 18 hours 3-base or 4-base T4 DNA ligase... 1 hour 3-base or 4-base T4 DNA ligase 37 °C 18 hours 3-base or 4-base Bbsl-HF®, 1x ...Ligase MM 37-16 °C cycling 4-base Bsal-HFv2® 37 °C Static 4-base Bsal-HFv2, 1x NEBridge...-16 °C cycling 4-base PaqCl, 1x NEBridge Ligase MM 37-16 °C cycling 4-base Sapl              ...overhangs with the same three nucleotides in a row; (4) no more than two nucleotides in the same position...used. Each of the tools works with either 3-base or 4-base overhangs, and with a variety of different enzymes...

  2. Who Gives a Tweet? 9 Facts About Scientists on Twitter

    Type
    Blog Post
    Published
    June 26, 2017, 1:31 p.m.
    ...scientific authorship in the United States (0.428). 4. The #1 (re)tweeted domain by scientists was Instagram...Sciencemag, Wiley, Cell and Scientific American Figure 4: Top Scientific Domains Tweeted by Biologists. 7. ...and analysis of scientists on Twitter. Plos One,12(4). doi:10.1371/journal.pone.0175368. PubMed PMID: 28399145...use Twitter professionally to network, learn more about alternative careers for scientists, and share cool....” This paper addresses the following questions about scientists on Twitter: who are they? What do they...

  3. Multiplex Genome Editing with CRISPR-Cpf1

    Type
    Blog Post
    Published
    May 9, 2017, 2:12 p.m.
    ....4% of HEK293T cells had edits at 4 of 4 targets when an array of 4 crRNAs, Cpf1, and a GFP tag were transfected...crRNAtracrRNACas9 crRNACpf1 Endonuclease size spCas9: ~4 kbsaCas9: ~3 kb ~3.9kb crRNA/gRNA length gRNA: ...transduction One vector expresses Cas9, eGFP, and up to 4 gRNAs Each gRNA requires its own promoter   Kabadi... this by showing that Cpf1 can cleave an array of 4 crRNAs in vitro and when expressed in 293 cells. This...Lastly, an AAV vector was used to multiplex edit 3-4 genes in a primary culture of mouse neurons and in...targets compared to Cas9.  Read on to learn more about Cpf1 multiplexing.  For an in-depth review of Cpf1... Addgene. If you have any questions or thoughts about Cpf1 multiplexing, leave them in the comments below...

  4. pCXLE toolkit: Efficient episomal plasmid-based method to reprogram peripheral blood cells to iPSCs

    Type
    Blog Post
    Published
    Dec. 14, 2017, 2:08 p.m.
    ...pCXLE-hUL 27080 L-MYC and LIN28 pCXLE-hOCT3/4-shp53-F 27077 OCT3/4 and shRNA against p53 pCXWB-EBNA1 37624...the expression of the reprogramming factors- OCT3/4, SOX2, KLF4, and C-MYC [1]. For the derivation of ...and remains a safety concern in the clinical field [4]. Three major reprogramming methods are considered...mixture of two vectors that encoded six factors - OCT3/4, SOX2, KLF4, C-MYC, LIN28 and SV40 large T antigen...PMID: 21243013. PubMed Central PMCID: PMC3193421. 4. Hacein-Bey-Abina, S., et al., LMO2-associated clonal...activated T cells and Sendai virus. Nat Protoc, 2012. 7(4): p. 718-28. PubMed PMID: 22422317. 6. Diecke, S.,...successfully to derive iPSCs from PBMCs. Most laboratories use Sendai virus to reprogram PBMCs efficiently...

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

    Type
    Blog Post
    Published
    March 7, 2017, 3:30 p.m.
    ... BRAF sites targeted by my specific gRNAs (Figure 4). The most difficult part of the assay was designing...surveyor assay itself was relatively easy. Figure 4: Detection of edited BRAF genomic loci in Cas9-expressing... cells transduced by the control GFP-gRNA (Figure 4, 5). When the gel was overexposed, I could even also...results. (Figure 5). The digestion product (Figure 4, lanes 0.3 % 3.0, Figure 5, lane 2) is faint, so comparing... comparing it to the undigested control (Figure 4, lane 0, Figure 5, lane 1) is what confirms it’s a real... control (GFP gRNA-targeted) BRAF sites (lanes 3, 4) were used as controls. Duplexes were generated from...amplification products (GFP-GFP duplex) as a control. 4. After optimizing the reaction and confirming the ...

  6. Top Requested Lentivirus and AAV of 2016

    Type
    Blog Post
    Published
    Jan. 6, 2017, 3:56 p.m.
    ...transduced area. Gene Ther. 2003 Feb;10(4):337-47. PubMed PMID: 12595892. 4. Sanjana NE, Shalem O, Zhang F. Improved...contains a double floxed human muscarinic receptor 4 (Gαi-coupled) DREADD fused to mCherry, all under the...feeding behavior in mice. J Clin Invest. 2011 Apr;121(4):1424-8. PubMed PMID: 21364278. PubMed Central PMCID...CRISPR screening. Nat Methods. 2014 Aug;11(8):783-4. PubMed PMID: 25075903. PubMed Central PMCID: PMC4486245...Resources on the Addgene Blog Learn More About Lentivirus Learn more About AAV Get the Scoop on Our Viral Service...

  7. Back to Bacteria: CRISPR gRNA Multiplexing Using tRNAs

    Type
    Blog Post
    Published
    June 2, 2015, 6:06 p.m.
    ...nuclease Csy4 from Pseudomonas aeruginosa to cleave 4 gRNAs separated by 28 base Csy4 recognition sites ...7 base poly(U)tail in tRNA-gRNA constructs or a 1-4 base tail from the second tRNA in the tRNA-gRNA-tRNA...al. constructed PTGs containing 8 gRNAs targeting 4 MAPK genes, again using rice protoplasts. Each pair...large deletions. They detected fragment deletion with 4-20% frequency at the four loci, indicating that Cas9..., Perez-Pinera P, Lu TK. Mol Cell. 2014 May 22;54(4):698-710. doi: 10.1016/j.molcel.2014.04.022. Epub ...with limited packaging capacity. Still confused about the various tRNA expression strategies? No worries... Addgene. If you have any questions or thoughts about this technology, leave them in the comments below...

  8. CRISPR 101: Multiplex Expression of gRNAs

    Type
    Blog Post
    Published
    Sept. 10, 2020, 11:45 a.m.
    ...plasmids are digested, unique overhangs (here, O1-4) adjacent to the cut sites “link” fragments together...plasmids: This plasmid set allows you to express 2-4 gRNAs, with four being the ideal number. First you...clone, from 2-7. For example, if you’re expressing 4 gRNAs, you’d use pX330A-1x4; for 6 gRNAs, you’d use... Plasmids: These plasmids allow you to assemble 2-4 gRNAs through Golden Gate or Gibson Assembly. gRNAs...all-in-one CRISPR/Cas9 vector system.” Scientific Reports 4 (2014): 5400. PubMed PMID: 24954249. PubMed Central...PMC4066266. Find plasmids from this paper at Addgene. 4. Albers, Joachim, et al. “A versatile modular vector... genetics.” Journal of Clinical Investigation 125(4) (2015):1603-19. PubMed PMID: 25751063. PubMed Central...

  9. 5 Tips for Troubleshooting Viral Transductions

    Type
    Blog Post
    Published
    Aug. 11, 2016, 2:23 p.m.
    ...ultracentrifugation (75,000 - 225,000 g for 1.5 – 4 hours at 4 °C). After ultracentrifugation, virus may appear...researchers prefer to store freshly harvested virus at 4 °C instead of storing it briefly at -80 °C. If troubleshooting...more concentrated virus solution can be obtained. 4. Enhancing virus-cell contact A variety of reagents...Physiology 291.2 (2006): C386-C392. PubMed PMID: 16510845. 4. Krajden, Mel, et al. "Effect of multiple freeze-thaw...grown at 30 °C instead of 37 °C. If you’re unsure about the integrity of your virus, perform a diagnostic...Additional Resources on the Addgene Blog Learn All about Adenovirus Associated Virus New to Virus? Check ...Check out Our Primer for Beginners! Learn All about Lentiviral Vectors Additional Resources on the Addgene.org...

  10. CRISPR Kinome Libraries Available: Pooled and Individual Plasmid Formats

    Type
    Blog Post
    Published
    Sept. 9, 2016, 2:43 p.m.
    ...two sets, each with 4 gRNAs per kinase, so screens can be performed with either 4 or 8 gRNAs per gene....the human kinome. These plasmids contain the same 3-4 gRNAs found in the first subset of the pooled library...numbers that’ll leave you, well, #sad. Learn More About CRISPR Pooled Library Screening Why the kinome?...

  11. Important Considerations When Using AAVs

    Type
    Blog Post
    Published
    June 13, 2017, 2:30 p.m.
    ..., hippocampus and cortex) within the mouse brain (4). An example from the cortex is outlined in the table...velocities of movement, distance traveled, etc.) (4). This has also been identified with AAV2 (5), AAV6... vector using a serotype with strong CNS tropism (4). If axonal transport is not desirable, transport...serotype with weak CNS penetrance (such as AAV2) (4). Otherwise, using a specific system for controlling...shortened (from ~4.8 kB [8] to ~2.3 kB [9]).     4. The Future What new types of research may be enabled...PMID: 24086725. PubMed Central PMCID: PMC3785459. 4. Castle, Michael J., et al. "Adeno-associated virus... Its Fruit Is Sweet The Future? I knew little about adeno-associated Viral Vectors (AAVs) before starting...

  12. Plasmids 101: Knockout/Knock-In Plasmids

    Type
    Blog Post
    Published
    Dec. 1, 2016, 3:30 p.m.
    ...only and leave exon 2 floxed, as seen in line 4 of Figure 4. Since the loxP sites are located in intronic...the targeted exon in the gene of interest (Figure 4). When recombination occurs, the gene still functions...has been placed into an intron.    Figure 4: A targeting vector to create a floxed allele for ...inserted GFP is expressed (knocked in). As seen in the above example, you can remove a floxed resistance gene...Get the Basics of Homology Directed Repair Read about Generating Knockouts with CRISPR through Non-Homologous...Expression Vectors Find Cre-Lox Plasmids Learn More about CRISPR  ...

  13. Technical Design of a Western Blot

    Type
    Blog Post
    Published
    Aug. 22, 2024, 1:15 p.m.
    ...250 kDa Different sizes 4–12% gradient 4–250 kDa Different sizes 4–20% gradient   Handcast...length of your assay. The most common options are 4 °C overnight or 1–2 hours at room temperature (RT)...choose to incubate their primary antibody overnight at 4 °C and their secondary antibody for 1–2 hrs at room...expensive, reduce protocol time, can be stored at 4 °C, and are very consistent gel-to-gel.  Ladder Whatever... stored at your bench during the procedure and at 4 °C when you’re not running a western. They are not...and will noticeably degrade after several months at 4 °C.  Pro tip! If you get a speckled image, your protein-based...If you’ve ever run a western blot, or thought about running one, you’ll know there’s a lot of choices...

  14. Podcast: A Malawian Professor's Path to Biotech Research

    Type
    Blog Post
    Published
    May 24, 2016, 2:30 p.m.
    ... and education 3:49-4:35 : Kingdom on opportunities to go into science in Malawi 4:36-5:46 : How Kingdom...more about Dr. Kwapata’s current work and thoughts on the need for more scientific collaboration across... connecting us with him. Learn more about our recent collaboration with Seeding Labs here. Special thanks...received an award from nonprofit and Addgene collaborator, Seeding Labs, for lab equipment that will help...Addgene Podcast.     Additional Resources Learn about Seeding Lab's Work with Lilongwe Univeristy of Agriculture...How You Can Contribute to the Addgene Blog Learn about Addgene Plasmid Donations for Rare Disease Research...

  15. New Podcast Segment: Hot Plasmids

    Type
    Blog Post
    Published
    May 17, 2017, 3:37 p.m.
    ... min) Hot Plasmids podcast that introduces you to 4 new plasmid technologies from one of our recent newsletters...The Human Kinase Domain Constructs Kit Hot Plasmid 4: The Yeast Prototrophy Kit from Markus Ralser's Lab...this new way to learn about plasmid technologies. If you have any comments about the Hot Plasmids Podcast...with these efforts, you’ll find new ways to learn about science careers, lab protocols, and, of course, ...each one, plug in some headphones to learn more about the latest and greatest plasmids through this podcast...

  16. Hot Plasmids: Spring 2025

    Type
    Blog Post
    Published
    June 3, 2025, 1:15 p.m.
    ...UTR sequence added to the genome (Figure 4). Figure 4: STITCHR consists of a nicking Cas9 fused...Glypicans are proteoglycans composed of a protein core, 2–4 glycosaminoglycans (GAG), and are (typically) tethered...Nature. doi: https://doi.org/10.1038/s41586-025-08877-4.  Two powerhouse tools combine for dynamic protein...mouse brain following retro-orbital AAV injection at 4 weeks. Cortical layers (L1-6) and scale bar are indicated...insert your genetic cargo of choice. After hearing about researchers' favorite features and limitations of...development and cancer. But there is still much to learn about their myriad functions and therapeutic potential...created a very quick and efficient way to learn more about gene regulation. Find the TurboCas plasmid here!...

  17. PiggyBac-ing Through the Genome Editing Field

    Type
    Blog Post
    Published
    May 31, 2016, 3:30 p.m.
    ...competent for excision, but defective for integration [4], the donor cassette is removed leaving behind no ... with the wild-type piggyBac™ transposase enzyme [4]. By using a donor plasmid containing a negative selection...Methods Mol. Biol., Apr. 2016. PubMed PMID: 27075976. 4.  X. Li, E. R. Burnight, A. L. Cooney, N. Malani, ...target locus modifications.,” Mol. Ther., vol. 18, no. 4, pp. 743–53, Apr. 2010. PubMed PMID: 20068556. PubMed... retroviral insertion sites.,” Nat. Protoc., vol. 4, no. 5, pp. 789–98, Jan. 2009. PubMed PMID: 19528954...transposase itself, please read on to learn more about this exciting technology from the folks at Transposagen...Additional Resources on the Addgene Blog Learn about the Sleeping Beauty Transposase Read up on Genome...

  18. Getting to Know Expansion Microscopy

    Type
    Blog Post
    Published
    June 20, 2023, 1:15 p.m.
    ...Depending on the protocol, samples can be expanded to ~4-10x linearly! This expansion physically separates ... cases, labels are added after this stage.  Step 4: Expand! (Typically by just adding water.)  Step 5...quite so much. Created with biorender.com Steps 2-4 are not found in standard IHC/ICC protocols and you...16:33–41. https://doi.org/10.1038/s41592-018-0219-4  Yu C-C (Jay), Barry NC, Wassie AT, et al (2020) Expansion...applications and there’s just not much to be done about that.  Second, expansion introduces distortions....

  19. Minigenomes - a Safe Way to Study Dangerous Viruses Like the Ebola Virus

    Type
    Blog Post
    Published
    March 17, 2016, 2:30 p.m.
    ...under the highest biosafety level conditions, BSL-4. In this regard, minigenome systems, such as the one...the EBOV replication cycle under BSL-2 conditions (4). What is a minigenome?   The EBOV genome is a nonsegmented... ratios result in significantly reduced activity (4). To ensure that you end up with appropriate plasmid...PMID: 21699921. PubMed Central PMCID: PMC3586226. 4. Mühlberger, Elke, et al. "Comparison of the transcription...Microbiology, National Emerging Infectious Disease Laboratories.  She is interested in genome replication and...

  20. Performing In Vivo CRISPR Screens Using the FITS Approach

    Type
    Blog Post
    Published
    May 2, 2024, 1:15 p.m.
    ... single gene KO and pooled screening approaches; (4) encodes a Thy1.1 selectable marker to facilitate ...Framework for in vivo T cell screens. J Exp Med, 221(4), e20230699. https://doi.org/10.1084/jem.20230699....faithfully model the complex spatial, cellular, metabolic, and cytokine microenvironment that exists in ...LaFleur is a Postdoctoral Fellow in Arlene Sharpe’s laboratory at Harvard Medical School and is interested in...
Showing: 1 - 20 of 340 results