... vs. SpCas9, see the table below and read on to learn more about GeoCas9.   SpCas9 GeoCas9 Size 1368...ranging from 25-80 °C. Activity is expressed as Kcleave (min-1). SpCas9 has maximum activity from 35-45...experiments tested the ability of GeoCas9 RNPs to cleave substrate when the reaction was carried out at ...either 37 °C or 4 °C. Substrate was then added and cleavage product analyzed on a gel. SpCas9 is stable when... 30% human plasma. The discovery of GeoCas9 clears the way for CRISPR/Cas9 genome editing in facultative...

... Addgene blog Learn about Gateway cloning Read about genome engineering in flies Learn more about controlling...HDVR (2X Hepatitis Delta Virus Ribozyme), a self-cleaving ribozyme. In the second-generation SPARC2 system...HDVR is placed upstream of the second attP site to cleave and inactivate any read-through transcripts of ... efficiency of recombination and preferentially leads to gene constructs where the stop cassette remains...

...engineering from ZFNs through TALENs and CRISPR-Cas nucleases, provided important context for the rest of the...reagents so freely with the academic community has clearly made a huge impact on this field. Advances in ...described a chimeric Cas9 that is improving the nuclease specificity dramatically and reducing off-target... service questions for nearly 7 years, and my colleagues and I have an internal motto for scientists who..., and I found myself extremely grateful to my colleague Jason for choosing Simon Singh's "The Code Book...

...Foundation, ABRC’s outreach program kicked off with the release of 20 education kits designed for use in K-12 and...ABRC database and ordering platform soon to be released. For individuals interested in ordering from ABRC...has many benefits. Engaging students in active learning through hands-on experiments allows them to develop... out opportunities to engage people in science learning. Whether visiting classrooms, hosting field trips....     Additional Resources on the Addgene Blog Learn how BioBuilder can bring Synthetic Biology into ...

...Resources at Addgene.org Learn about Popular Plasmids in the Repository Learn about New Plasmid Technologies... relatively low efficiency of biotinylation, at least in vitro. Our solution was to exploit the best of...proteins that lacked TEV. Although we were unable to cleave TEV within the protein we tested, it may be possible...plasmid in NEB T7 SHuffle bacteria (C3029J). We are clearly NEB “fanboys” as the bacteria significantly enhance...

...structure-guided direct evolution, Nathan Shaner and colleagues from The Scintillon Institute, Allele Biotechnology...Clavularia sp. protein CFP484. Hui-wang Ai and colleagues from the University of Alberta and University... in a 2.6-fold increase in signal relative to mCerulean imaged with a standard CFP set. mTFP1 can be used...would like to share with the scientific community, please deposit the plasmids at Addgene! References Ai... to the Addgene blog Resources on Addgene.org Learn about the Michael Davidson fluorescent protein collection...

...Additional resources on Addgene.org Learn about Addgene's viral service Learn about viral production Find viral...incorporate into their own labs. This week we are pleased to announce the submission of the results of our... cells (Vandenberghe, 2010). Rather than being released into the cell culture supernatant, the viral particles..., Luk H., et al. "Efficient serotype-dependent release of functional vector into the culture medium during...

...ever like to write about a recent plasmid deposit please sign up here.   Listen this episode of Hot Plasmids...contain a smaller RNA tag, are less prone to photobleaching, and achieve better signal in vivo as compared...increasing the brightness. Arne Fabritius and colleagues started with the FP mNeptune684 (originally described... rapid quantification of base-editing and Cas9 cleavage activity Article contributed by Angela Kaczmarczyk...show that the ACE reporter can also quantify Cas9 cleavage, thus exhibiting its versatile utility in quantifying...

...Figure 3: When a bison wanders into the campsite, leave your samples behind! Paul and Anastasia (Potanina...hyperthermostable Cas9 This year it was wonderful to learn that one project led by postdoc Dr. Stephanie Schmidt... IgnaviCas9 is a hyperthermostable Cas9 CRISPR nuclease that is active up to 100°C.  The IgnaviCas9 protein...for long time periods and workflows where CRISPR nuclease activity is desired and time or protocol steps...useful. Additional resources on the Addgene blog Learn about other Cas proteins Read all of our CRISPR ...

...excitation and emission spectra. Both spectra possess clear maxima where excitation and emission are the most... 500 nm and a global maximum at around 560 nm, leading to different levels of brightness, depending on...check out our blog post on color compensation to learn more about them. Alright, let's build your first...cell culture, the transduction process itself also leads to some cell death. As a result, there is a high...   Conclusion You made it! In this post, you learned about the principles behind fluorophores and their...this process creates visible colors. Next, you learned about how fluorophores can be classified into different...

...with a microbe, to trigger immune cascades that clear these infections. Anything that generates an antibody...immune system is referred to as an antigen. You can learn more about antibodies in our Antibodies 101 animation...isolate cells expressing this protein. Read on to learn more about antibodies and how to use them in the...produce is not necessarily what you are getting. Learn more about monoclonal antibodies here. Recombinant...simultaneously as well as compare between samples. Learn more about flow cytometry. Cell and tissue labeling...antibodies, the Developmental Studies Hybridoma Bank Learn more about nanobodies Watch our Antibodies 101 animation...

...attempted to study cancer from every possible angle to leave no stone unturned, but cancer has been producing...between tumors and the immune system We’ll begin by learning how the immune system naturally interacts with...Through interactions with the immune system, tumors “learn” how to escape it. In this process, cancer cells...tolerance-producing antigens enunciated above. Finally, signals released from the microenvironment help create tolerance...enter the clinic, physicians and researchers are learning much about how treatment regimens can be improved...PMC3433802. Additional Resources on the Addgene Blog Learn About Our Cancer Collection Pages Plasmid Grant ...

...contributed by guest blogger Fredrik Wermeling, leader of a research group at the Centrum for Molecular...design gRNAs for all the purposes discussed above. Please see videos and text on the website for more detailed...have been “green listed” by several algorithms. Please read the “Detailed Information” text linked to ...and libraries using strategies related to other nucleases as well as inhibitory and activating approaches...PMC5292764. Additional Resources on the Addgene Blog Learn about CRISPR Screens How to Design Your gRNA Tips...for a 1st CRISPR User Resources on Addgene.org Learn More About CRISPR Find CRISPR Screening Libraries...

...than 75% photobleaching after just 1 minute of exposure. MG-2I-dL5** experienced minor bleaching after longer...ablate cells in culture and in vivo.  Read on to learn more about this killer illumination technique! ...real time.  In both cases, dL5** was tagged with mCerulean3 (mCer3) for visualization and expressed from ...Sign Up to Receive the Fluorescent Protein eBook Learn about Small Molecule Fluorophores Resources on ...

...ride is a great activity for both exercise and clearing your mind. And because the outdoors is so huge...events. Download Addgene’s Science Career guide to learn more about career paths and things you can do to...Plan out meals and snacks ahead of time.  Find learning activities for kids. For example, Skype a Scientist...collected into figures for papers or presentations. Learn a new skill for analyzing and visualizing your data...introductions and literature reviews could make the months leading up to your defense less stressful. Think about...

...updates to iGluSnFR and SF-iGluSnFR have made it clear that it’s time to update our iGluSnFR post! Here...give researchers the ability to monitor glutamate release from neurons and other brain cells in vivo. Glutamate...protein binds to glutamate, conformational changes lead to an increase in fluorescence intensity from the...differentiating between synaptic vs. extrasynaptic glutamate release. Building on the previous generation of the SF-...Resources Additional Resources on the Addgene Blog Learn about other Fluorescent Biosensors A Practical Approach...

...incorporation of a 3’-5’ exonuclease in the reaction mixture. The exonuclease removes mismatched 3’ overhangs...guide the joining reaction. BioBrick Assembly will leave scar regions at the site of fragment joining, but...still creating scarless plasmid inserts.  Want to Learn More about DNA Assembly? Check Out Our Plasmids ...      Additional Resources on the Addgene Blog Learn More about Gateway Cloning Gibson Assembly and Other...

... of the mRNA, leaving only a red FP bound to the reporter. Because it contains a nuclear localization ...cell. When cellular mRNAs are digested with ribonucleases, ribosomes protect the segments of mRNA to which...occur. For a TRICK reporter mRNA this displacement leads to a change in the fluorescent signal: untranslated...Addgene. Additional Resources the Addgene Blog Learn More about SunTag Catch up on Your GFP Background...

...complex processes can’t be overstated, we can also learn a lot from Nature’s simpler solutions to transmitting..., and they can rupture E. coli spheroplasts to release encapsulated proteins. Video by ACS. For a more...appealing target for bioengineering, but it wasn’t clear whether they could 1) be decorated with proteins...Tunable Protein Piston That Breaks Membranes to Release Encapsulated Cargo. ACS Synth. Biol. (2016). doi...

...libraries Learn about other members of the Addgene Advisory Board Additional Resources Learn a little...post-doc, Doench worked on using RNAi screens to learn more about cancer biology in Ed Harlow’s group at...and “with CRISPR, just the results are so much cleaner. The time frame from doing your primary screen ...Institute here Read other blog posts about CRISPR Learn more about the Broad Institute and the Platform ...