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We narrowed to 727 results for: cat.1

Showing: 391 - 420 of 727 results

  1. Hot Plasmids - May 2022

    Type
    Blog Post
    ... Zebrafish) (Xiong, et al., 2021).    Fig. 1: Schematic showing the workflow for the BLITZ system...Biol. 2022. https://doi.org/10.1038/s41556-021-00836-1 Visual barcodes for live cell clonal multiplexing...of interest and, for in vivo work, generate a dedicated transgenic line. Using zebrafish as a model organism...off-target mutations. These plasmids contain catalytically inactive dCas9 and microbial SSAPs (dCas9-SSAP...clonal multiplexing of cells in live imaging application. Each visual barcode consists of a fluorescent...transcription response element (TRE) or kinase translocation reporters (KTRs). Find visual barcode plasmids...

  2. How to Design Your gRNA for CRISPR Genome Editing

    Type
    Blog Post
    ...non-functional allele. For a 1 kilobase gene, since potential target sites occur ~1 in every 8 nucleotides,...770–788 . https://doi.org/10.1038/s41576-018-0059-1 Veres A, Gosis BS, Ding Q, Collins R, Ragavendran ...the optimal location. It is also important to have good information on the exact location of the TSS. ...appreciable levels of modification. Whole-genome sequencing of cells modified by CRISPR indicates that the consequences...specific base edit, or modulation of gene expression. Location and sequence are important considerations for ... gRNAs. For indels, it's not so important what location in the gene you target, but it is important that...fewer sequences to choose from). Finally, for HDR, location is much more important because you have to target...

  3. Plasmids 101: Inducible Promoters

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    Blog Post
    ...Hsp70, the heat shock releases heat shock factor 1 (HSF-1), which subsequently binds to heat shock elements...prokaryotic promoter often used for bacterial protein purification. When arabinose is absent, regulatory protein...

  4. CRISPR/Cas9 FAQs Answered!

    Type
    Blog Post
    ...new data). For general transfection, you could use 1 µl of 10 µM oligo stock for each well on 24-well plate...A10: The most we've tried to insert so far has been 1 kb. We used homology arms that were 800 bp long. Q11...able to cut). Clones without insertion will show a ~1 kb and ~7.5 kb fragment (both BbsI and AgeI will be...Mammalian Cells. Molecular and Cellular Biology, 18(1), 93–101. https://doi.org/10.1128/mcb.18.1.93  Ran...with the newest CRISPR technologies and their applications can be exhausting. A quick search for "CRISPR... the target site sequence (protospacer+PAM) is located? If there are other PAMs in the region, will they...puromycin selection. You could then confirm this purification by performing a genotyping assay (such as Sanger...

  5. All in a Twist: dsRNA

    Type
    Blog Post
    ...versions with toxicity to many organisms. Fig. 1: Exogenous sources of dsRNA Cellular sources of... 286-301 (2022). doi.org/10.1038/s41580-021-00430-1 More resources on the Addgene blog CRISPR 101: RNA...generate dsRNA as part of their infection and/or replication process within the host. dsRNA viruses are inherently...generate dsRNA intermediates as part of the viral replication process. dsDNA viruses yield dsRNA through a ...

  6. Gendered Innovations: Why Does Sex of the Cell Matter?

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    Blog Post
    ...healthcare system was $35.2 billion (or $140 per $1 invested in the trial). The goal of gendered innovations...is the goal of Gendered Innovations. This project: 1) develops state-of-the-art methods of sex and gender.... To remedy this, Shah et al. provide a table indicating the sex of the most commonly used cell lines ... work to be done! Researchers need to learn sophisticated methods of sex and gender analysis. Universities...

  7. Deep Dive: Statistical Tests (Comparisons)

    Type
    Blog Post
    ... closely data points fall to each other (see Fig. 1). Alpha values – numerical risk allowance in a ...statistical test assumes your data follows. Fig. 1: Examples of comparable groups with and without equal...values from smallest to largest and ranking them (1 being the smallest value). It calculates the cut off...tests  Statistical tests fall into two general categories: parametric and non-parametric. In this blog ... exist somewhere within your data; it does not indicate which groups the differences exist between. For...that matter. However, this is oftentimes an oversimplification. If your independent variables are affecting...

  8. 5 Steps to Writing Grant Proposals for Grad Students

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    Blog Post
    ... as a Professor. Step 1: Your PI forwards you a Request For Applications (RFA), what next? First,...proposal typically lists the deliverables, explains 1-2 potential pitfalls of the intended protocol, and...the grant application process. When my PhD advisor first asked me to write a grant application due the ...in a previously funded grant application to see how funds were allocated. Step 3: Get feedback from your...the next round of applications. Step 5: Keep writing! Each written grant application builds upon the ... project grant application" by the NIH Still want more information on the application process? What types...advisor alone applies for these competitive grant applications; however, in some cases, your advisor may choose...

  9. CRISPRainbow and Genome Visualization

    Type
    Blog Post
    ...all the colors of the CRISPRainbow. Figure 1: CRISPRainbow was used to track the localization of...is available as a kit from Addgene.   References 1. Chen, Baohui, et al. “Dynamic Imaging of Genomic ... every genomic locus, and to do so in vivo. Catalytically dead Cas9 (dCas9) cannot induce gene editing...Cas9 ortholog, and the target sequence must be located adjacent to that PAM. To scale up fluorescent CRISPR...this rainbow of colors, Ma et al. performed sophisticated tracking of multiple chromosomal loci in live... do not induce genome editing. If used with catalytically active Cas9, the short, CRISPRainbow gRNA will... PMCID: PMC3918502. Find plasmids from this publication at Addgene. 2. Ma, Hanhui, et al. “Multicolor...

  10. Plasmids 101: Golden Gate Cloning

    Type
    Blog Post
    ... the original plasmid (Option 1) or added using PCR-based amplification (Option 2). Advantages of Golden...Methods Mol Biol. 2014;1116:119-31. doi: 10.1007/978-1-62703-764-8_9. PubMed. More Resources at Addgene:  ... work around this is to "domesticate" your fragment: PCR-based amplification can be used to create silent...and exciting applications, this system is an incredibly powerful tool for cloning complicated constructs...Type IIS sites (such as BsaI or BbsI), that are located on the outside of the cleavage site. As a result...restriction sites that may not be amenable to "domestication", you might want to consider using an alternative...high-efficiency step. References: Sequential amplification of cloned DNA as tandem multimers using class-IIS...

  11. New FLIS Plasmid (April Fools 2022)

    Type
    Blog Post
    ... plasmid-based system for locating and containing escaped flies. Nature 4:1-22. https://doi.org/4.01.2022...Additional resources on the Addgene blog:  Flies 101: Catching flies in the lab References Abril, U.N.O., et....

  12. Hot Plasmids and Viral Preps - March 2021

    Type
    Blog Post
    ...and accurately in a “single pot.’   Figure 1: Single-Pot Assembly: a) Graphic depiction of chromatin...AID system relies on a transport inhibitor response 1 (TIR1) expressed under a desired promoter with a specific...2021.   The potential tools fall into two categories: probes and editors. A probe array may consist...and should be varied as it can have important implications for the fusion protein effectivity), and a fluorescent...to visualize DNA, an editor targets DNA (via catalytically inactive dCas9) to modify chromatin. A fluorescent...both spatiotemporal protein degradation and quantification of this depletion using imaging-based measurements...BioTag, which can be used in protein affinity purification studies.    Duong et al., Genetics 2021. https...

  13. The Future of Research Symposium Boston 2015

    Type
    Blog Post
    ...all of its successes, has many deep-rooted flaws (1). As we discovered through the four workshops at the... Follow David on Twitter @driglar.     References 1. Alberts, Bruce, et al. "Rescuing US biomedical research...University of Massachusetts Medical School. Graduate education programs are also beginning to address these issues...The panel discussion on Friday morning will be dedicated to Academic data and the Labor Market. On this...Sinche is a STEM career counselor, Director of Education for the Jackson Laboratory for Genomic Medicine...disease.  Outside the lab, he is interested in communicating science and in the intersection between science...

  14. Viral Vectors 101: Gamma-Retroviral Packaging Systems

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    Blog Post
    ...additional flexibility in pseudotyping.   Figure 1: Genome of a prototypical gamma-retrovirus.    ...an SV40 promoter, and Neomycin selection.   Table 1: Gamma-retrovirus glossary Plasmid Type Element... be used interchangeably. Are γ-retroviruses replication incompetent? Yes, as with lentiviruses, the genes...

  15. Plasmids 101: Screening Strategies Used in Plasmid Cloning

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    Blog Post
    ...screening blog post for more information.   Figure 1: Result of a blue-white screen. Image from Stefan ...Because Sanger sequencing can usually sequence just 1 kb of DNA, custom primers are especially helpful in... of the entire plasmid. Please note that NGS verification is more time-intensive than Sanger sequencing...

  16. CRISPR 101: Non-Homologous End Joining

    Type
    Blog Post
    ...Last updated Sept 1, 2020. This post was contributed by David Wyatt and Dale Ramsden, UNC at Chapel Hill...the course of repair by NHEJ are typically small (1-10 bp) but extremely heterogeneous. There is consequently...Giovannangeli C, Jasin M, Brunet E (2014) Chromosomal Translocations in Human Cells Are Generated by Canonical Nonhomologous...

  17. Plasmids 101: The protein expression toolbox

    Type
    Blog Post
    ...introduced, they can deplete endogenous protein levels for 1-3 days. There’s also a CRISPR for that! CRISPR interference...more about them on our CRISPRa blog! Fig. 1 CRISPR activator and repressor mechanisms Want...interference (CRISPRi) targets a catalytically dead Cas9 fused to a transcriptional repressor to your ...

  18. Troubleshooting Your Plasmid Cloning Experiment

    Type
    Blog Post
    ...are working with is correct. It's also best to use 1-2 ug of the vector for digestion. PCR Design primers...plasmid; a starting point could be a molar ratio of 1:2 (plasmid:insert), but this parameter has to be optimized...constructs. His work was acknowledged in hundreds of publications including Cell, Nature, and other journals. ...

  19. CRISPR 101: Cas9 vs. The Other Cas(s)

    Type
    Blog Post
    ...main differences between Cas7-11 and Cas13 are 1). non-specific RNA cleavage and 2). cell toxicity. Cas7... targeting single stranded RNA.   Table 1: Fast facts on Cas family proteins and genome editing...a defined site to form a clean DNA break, Cas3 locates its target sequence and initiates single stranded... out large regions of DNA and has anti-viral application potential as a result. Fast facts – PAM requirement... has been proposed to be useful for the same applications (RNA knockdown, RNA editing, etc.). The main...

  20. New Videos: Addgene Lab Tips

    Type
    Blog Post
    ...the lab running like a well-oiled machine. Lab tip #1: Glove trick Putting on lab gloves can sometimes be...With barcode tracking, you will always be able to locate materials quickly and easily. Learn more about ...Additional Resources on the Addgene Blog New Educational Resource: The Addgene Videos Page 5 Science Rap...

  21. How to Prepare for an Industry Interview

    Type
    Blog Post
    ...company culture.  It’s also a good idea to prepare 1-2 unique questions for each interviewer by looking...to take notes after each interview.   Tailor your 1-hour presentation to your audience Start by asking...accomplishments and demonstrate your science communication skills. Know your audience. Connect your prior...codes for interviews. You can also ask HR for clarification on the dress code for the interview. Greet each...

  22. 6 Tips for Grant Writing

    Type
    Blog Post
    ...writing and hopefully increase your success rate. 1. Submit your grant early! First off, and perhaps the...involved so it’s important not to throw in the towel if 1 or 2 funding bodies say no. If possible, get feedback...Series 4. Become a science communicator More and more fellowship applications are asking for a 200-300 ...good quality application. Even following this advice, only about 50% of my grant applications have been ...do need to be able to research and write your application, then give it to someone else to read and / or...it). It’s easy for reviewers to spot when an application is rushed (whether the idea isn’t properly researched...researched / developed or the application is full of typos) and these are unlikely to get past the initial...

  23. Adapting Toehold Switches to Detect Zika Virus

    Type
    Blog Post
    ...self-assembly for applications in health, energy, and sustainability.   References 1. Pardee, Keith, et...detects samples at clinically relevant concentrations (1-3 fM; 1x106-fold lower than before), and our combined... journal Cell just six weeks later. The Zika publication timeline I think we can speak for all the authors...research, but the urgent need for the test did catalyze our resolve to get the work done as quickly as... dozen versions of the toehold switches and amplification primers. This helped make sure that we had a...this includes primer directed isothermal RNA amplification, new toehold switch designs, CRISPR/Cas9-mediated...our electronic reader that provides portable quantification of diagnostic results.  Our approach now detects...

  24. Simple CRISPR-based Epigenetic Editing: dCas9-directed DNA Demethylation

    Type
    Blog Post
    ...: it’s dCas9… with no modifications). Ready to get started?   Figure 1: Break free from the ball...?  Dividing cells depend on DNA methyltransferase 1 (DNMT1) to copy methylation marks from parental to... when DNMT1 methylates the genome during DNA replication, it doesn’t methylate sites blocked by dCas9....for dCas9 to interfere with DNMT1 even through replication. The dCas9 also needs to be removeable, otherwise...vivo using dCas9–peptide repeat and scFv–TET1 catalytic domain fusions. Nat Biotechnol 34, 1060–1065. ...

  26. Antibodies 101: Fab Fragments

    Type
    Blog Post
    ...Fragment? Consider the classic IgG antibody (Figure 1): it is composed of four peptide chains (two identical...that actually bind antigens.      Figure 1: IgG antibody structure with important features labeled...fragments described above can be used for similar applications and have similar advantages, though with some...

  27. Antibodies 101: Stripping and Reprobing Western Blots

    Type
    Blog Post
    ...Glycine-HCl 25–200 mM SDS 0.1%–1% (w/v) Tween 20 (optional) 1% (v/v) HCl Adjust to pH 2.0... tool to identify a protein of interest in a complicated solution like cell lysate. But they can be a ...

  29. 5 Science Rap Videos & Their Creator: Tom McFadden

    Type
    Blog Post
    ...forms of science communication. Click here to listen to the interview with Tom McFadden!   1. Rosalind Franklin... from Addgene.      Check out more science education tools like Verse Two and Fill in the Planck at...with us. Tom McFadden is a YouTuber and science educator looking for creative ways to implement the next...next generation science standards and communicate science. Learn more about Tom on his youtube channel ...Community Check out All of Our Podcasts Find Great Educational Content in our Plasmids 101 Series Additional...

  30. When Fidelity Matters: A frank discussion about ligase fidelity

    Type
    Blog Post
    ...phosphorylated DNA termini to 3 ́-OH DNA termini (1-4). In vitro, ligases (notably T4 DNA ligase) are ... enzymology of DNA repair enzymes.     References 1. Shuman, Stewart, and Christopher D. Lima. "The polynucleotide...thermostable DNA ligase-encoding gene." Gene 109.1 (1991): 1-11. PubMed PMID: 1756968. 7. Lauer, G., et al. "Cloning...or DNA base pair mismatches (8-11). For some applications, there cannot be any end-joining activity, and...regions. Why use a high fidelity ligase? Numerous applications have been developed to take advantage of the...Taq and other thermostable DNA ligases. These applications detect specific nucleotide sequences with high...rounds of melting and annealing allows linear amplification of the probe ligation product. With the right...
Showing: 391 - 420 of 727 results