...lucky" and find a great gRNA with a proximal PAM right where you need it, the odds of this occurring are...reliability. Notably, the SpRY variant didn’t display higher off-target effects than SpCas9, a pleasant surprise...stranded cleavage and base editing applications, with high editing capacity in both contexts for NRN PAMs and...

...host) as well as the introduction of the cancer-fighting component, CAR. Clinical trials are underway to...allele specifically. Early-stage trials showed a high success rate in clearing misfolded protein levels...Regarding effectiveness, if the editing frequency isn’t high enough to make a physiological difference, then ...

...ER lumen reaches the sub-millimolar range, much higher than in the cytosol. Thus, for the ER you would...subsequent change in fluorescence only occurs at high calcium concentrations. This will reduce noise and...simultaneous (1) user-controlled illumination with violet light and (2) exposure to increased intracellular calcium...

...recombinase systems. Picking Ligands Picking the right ligand is also important for building a chemogenetic...pharmacologically selective effector molecules (PSEMs). Higher potency means lower drug doses, which decreases... CH, Coward P, Degtyarev MY, Lee EK, Kwa AT, Hennighausen L, Bujard H, Fishman GI, Conklin BR (1999) Conditional...

...company. This might seem like a no-brainer, but it happens more frequently than you might think. The Addgene... crucial information about a product is printed right on the tube itself! Is the answer to your question...

...colonies by (carefully) holding plates up to the light, where they appear as opaque dots (colonies should...the agarose onto the recipient plate at just the right temperature. If the agarose solution is too hot,...depending on the level of endogenous expression. We highly recommend the use of an electronic pipettor with...

... of interest thus represents a straightforward means of exerting tight control over Cas9 nuclease activity...different genetic loci within a single cell. One might imagine transfecting cells with both the nuclease-positive...

...primers.  Extension- The temperature is then raised slightly to Taq polymerase’s ideal temperature (70-75oC...which can be time consuming and challenging to get right. ddPCR uses a water-oil emulsion droplet technology...those stated above. Multiplex PCR is also used in high throughput SNP genotyping, gene deletion analysis...

...something silly like “See there, arginine 53 really is right there, at the end of helix 3…just like we said it...are useful. While a physical model of a protein might represent one aspect of the protein very well, it...while and you will wake up in the middle of the night dreaming about the conformational gymnastics that...

... Sometimes people don’t know how to start. They might want to use our promoters and terminators, but then...coding sequence to the MoClo system. It’s quite straightforward really and we’ll be producing an instructional...chances of doing something amazing will be equally high as for those who go to work in cancer biology. ...

...to make the most of your time and sequence! You might also enjoy seeing how our quality control process...alignment results. The default setting is “Optimize for Highly similar sequences (megablast)”, which is very fast...nucleotide sequences, proteins often have repeated or highly homologous regions, which by default are ignored...

...a collection of promises to not assert property rights for particular engineered genetic sequences; but...metabolite in a cell. The only way to do that at scale right now is to grow up a bunch of cells, sacrifice them...chemicals, to make more medicine. The parts we have right now are well adapted to work in prokaryotic bacteria...

...available Not commercially available Small but Mighty: Advantages of Nanobodies Although antibodies have...target antigen to about 2 nm, which can lead to higher resolution for super-resolution microscopy. Nanobodies... two questions, refer to this table to find the right nanobody for your work. Species Antibody Subclass...

...standard curve for qPCR can be challenging to get right. Here at Addgene, we use a linearized plasmid standard...very few positive droplets. In the image to the right, there is approximately 1 positive copy per microliter...droplet generator to the PCR plate. Maintaining a high droplet count is important for calculating the concentration...

...trainees’ very way of thinking too. Albert Einstein might have put it best when he stated, “the value of an...pathways (precisely the type of exercise Einstein might have resented!). In 1997, Joseph Provost, then a...like an exciting opportunity for your classroom, I highly recommend you try implementing one. It's likely...

... know the basic principles of IF, we’ll take a slightly deeper dive into those steps and provide you with...commonly used cross-linker for IF. However, you might come across a couple other terms that can cause ...especially if left on for too long or used at too high a concentration.      Figure 2: Examples...

...A) Streamlined VchINT expression constructs. (B) High integration efficiencies achieved with VchINT, at...mini-transposon Transform plasmid(s) into cells and plate overnight Screen colonies phenotypically or by PCR/sequencing...Sternberg SH (2020) CRISPR RNA-guided integrases for high-efficiency, multiplexed bacterial genome engineering...

... off-target effects. Cas9 RNPs are detectable at high levels shortly after transfection, and are quickly...efficiency, while biolistic bombardment, which can have high efficiency, requires specialized equipment, such...microbiology.     References 1. Chen, Sean, et al. "Highly efficient mouse genome editing by CRISPR ribonucleoprotein...

...interested audiences quickly and concisely – video might be part of the answer. Problems with written publications...TedTalks are very popular, though they focus mostly on highly innovative ideas rather than on the day-to-day ...that provides links to the videos, keeping the copyright and control over the videos with the researcher...

... #MolluscMonday: #Crepidula atrasolea juvenile, light microscope with polarizing filter. Development is...an extreme number of nerve cells and an unusually high level of RNA recoding in the brain. RNA editing ...DNA in the eggs to get transgenic animals. They might be immortal. Manihot esculenta (Blake Meyers) Cassava...

...million dollar sales pitch. Given this large sum, one might expect that applicants would work for months to ...research? This is the central idea. It should be at a high level, broadly understandable to scientists outside...engaged by your presentation, then you’re on the right track. Furthermore, practice getting back to your...

...data inside the ELN is reproducible, and only the highly restricted number of people who can access the ...practical to move all this data in bulk into a lightweight tool like an ELN. It is nevertheless critically...

...particular gene. Furthermore, a target sequence may have high homology elsewhere in the genome. These off-target...Sontheimer EJ (2018) NmeCas9 is an intrinsically high-fidelity genome-editing platform. Genome Biol 19...Evolved Cas9 variants with broad PAM compatibility and high DNA specificity. Nature 556:57–63 . https://doi....

...depositing lab. (For our more popular libraries, you might receive a similarly-sized aliquot from an amplification...ll want to infect your cells with the virus–the right amount of virus, that is. Ideally, each cell will...Konermann S, Gootenberg JS, Abudayyeh OO, Platt RJ, Brigham MD, Sanjana NE, Zhang F. Genome-scale CRISPR-Cas9...

...recombination at a frequency three orders of magnitude higher than identical targeting vectors delivered through...Other investigators who had been working to develop high efficiency gene targeting approaches improved on...repair, it has proven more challenging to develop high efficiency CRISPR-based approaches for the generation...

.... This over-reliance on the perfect story for a “high impact” publication can lead to the publication ...reproducibility issues in the biological sciences. I’d highly recommend listening to the panel discussion to ...Scientific Figures into Dashboards and Share Your Insights Who gives a tweet? 9 facts about scientists on...

...important that your gRNA sequence is designed to be highly active and reduce off targets. For CRISPRa and ...consuming process and should not be undertaken lightly! Indeed, truly achieving the gold standard requires...genetic screens using genome-wide libraries have shown high concordance between different sequences targeting...

... LITE system, which uses light to regulate transcription, also includes light-regulated histone methyltransferases... CA (2017) CRISPR–Cas9 epigenome editing enables high-throughput screening for functional regulatory elements...

...may be a good choice for targeting neurons, with high transduction frequencies in the central nervous ... our in-stock viral prep service.  AAV8 AAV8 has high transduction efficiency in multiple tissues, rapid...AAV9 AAV9 has relatively broad tissue tropism and high transduction efficiency, and is an excellent choice...

...using them. Indeed, the future of mouse work is bright as it is now easier than ever to manipulate the...technology, CRISPR, that is simple in concept, straightforward to use, and robust in performance. In its natural...BL, Xavier RJ, Root DE (2014) Rational design of highly active sgRNAs for CRISPR-Cas9–mediated gene inactivation...

.... PubMed PMID: 18845712. 9. Warren, L., et al., Highly efficient reprogramming to pluripotency and directed...PubMed PMID: 26189426. 15. Si-Tayeb, K., et al., Highly efficient generation of human hepatocyte-like cells...Central PMCID: PMC2946078. 16. Zhang, D., et al., Highly efficient differentiation of human ES cells and...

...existing content. Access the CRISPR Guide here! Highlights include a much-anticipated prime editing section...

... an antibody that works, we highly recommend “Chapter 3: Finding the Right Antibody for Your Experiment...

...protein for controlling molecular processes with light. Listen to Our Segement on PhoCl Segment  Hot Plasmid...Plasmid 4: pPSU Plasmids for generating DNA molecular weight markers.  Listen to the Segement on the pPSU Plasmids...

...generating knockouts using CRISPR and Wednesday will highlight some Cas9 variants. Next, we’ll look at various...

...resources! Ready to sing along with this year's lineup? Right! We're ready to, so let's get started!  On the...sequences🎵On the 2nd day of CRISPR, Addgene gave to me: High-fidelity Cas13 variants for targeted RNA degradation...

...adorned with a bioluminescent tree topper, tube “lights,” and autoclave tape garland. Thanks @ribosholmes... bit of everything: frog snowflakes, ornaments, lights, and took a group photo wearing holiday sweaters...

...from the first generation of base editors to the eighth generation (and dual base editors, too!).  On the...gave to me: CRISPR methods for bacteria.  On the eighth day of CRISPR, Addgene gave to me: anti-CRISPR ...

... In each short episode of this series, we’ll highlight one tip that we’ve learned in the lab that will...

...to make inexpensive 100 bp or 1 kb DNA molecular weight ladders were recently deposited with Addgene. A... The pPSU Plasmids for Generating DNA Molecular Weight Markers. Scientific Reports,7(1). doi:10.1038/s41598...