...techniques differ in important ways, but can also complement one another by providing both broad and specific... predict off-target effects of sgRNAs using computational methods. They identify possible off-target sites...the clinic. NGS-based methods provide the most complete information profile regarding putative off-target... with in silico prediction to create a more comprehensive list of off-target editing events. Two of the...tissue from whole animal models, no exogenous component required (e.g. dsODN), doesn't require high read...Vouillot, Léna, Aurore Thélie, and Nicolas Pollet. "Comparison of T7E1 and surveyor mismatch cleavage assays...the limit of detection for Sanger sequencing: a comparison of methodologies for KRAS variant detection."...

...minutes of inactivating the CASANOVA complex, fluorescent CRISPR complexes were observed binding telomeric ...using blue light (Bubeck et al., 2018). They accomplished this by inserting a photo-sensitive LOV2 domain...indicator podcast segment The neurons in the brain compute using electrical impulses to coordinate emotions...sensitivity, responds quickly, is photostable, and is compatible with optogenetic control. The researchers also... the “2in1” vector cloning system - a Gateway compatible approach to simultaneously clone two genes-of-interest...2015) or ratiometric Bimolecular Fluorescence Complementation (rBiFC, Grefen and Blatt, 2012). The 2in1 Plasmid...

... The Association for Creatine Deficiencies: Compensating for SLC6A8 mutations in Creatine Transporter...channels typically expressed on other cell types can compensate for or bypass the dysfunction of another solute...cells to identify any solute channels that can compensate for the defect caused by SLC6A8 deficiency. “...where one of the X chromosomes is partially or completely missing in women. Bicuspid aortic valve (an aortic...with an emphasis on oligomerization, signalling complex assembly, and localization to the nucleus,” says...at McGill University.  These protein-receptor complexes consist of many parts that work together on the...gene, which codes for G𝛃1 subunits of G protein complexes, lead to a rare type of developmental delay, and...

...preserves RNA structure and naturally occurring complexes present in the cell, it can also introduce non-physiological...interest. See figure 1 below for details on the key components of RaPID. Ramanathan et al. also tweaked BirA... BirA* homolog found in B. subtilis. BASU was  compared head-to-head with the E. coli and BioID2 BirA*...streptavidin signal (~250-fold enrichment) when compared to a scrambled RNA motif control). BioID2 had ...with BASU still generated a >30-fold enrichment compared to E. coli’s BirA*. Faster labeling kinetics also...HHCS-associated sequences had lower IREB2 binding enrichment compared to wild-type sequences and that loss of binding...Russell, P., & Guttman, M. (2014). Methods for comprehensive experimental identification of RNA-protein interactions...

... SCF E3 ubiquitin ligase complex. In the presence of auxin, this TIR1 complex binds to the degron sequence...depiction of chromatin within a nucleus -> DNA compaction around histones -> Examples of histone-modifying...included for visualization purposes, but the key component of the editing assemblies are sensor-actuators...The tool-kit consists of highly useful vectors compatible with Gibson and Saptrap cloning systems to generate...Resources page.  Plasmids for delivering CRISPRi components by a single AAV vector are now available from... developed by Taekjip Ha’s lab where Cas9-gRNA complex binds the target DNA without cleavage until it'...

...-Vvd showed substantially improved activity as compared to other recombinase/photodimer pairs. Created.... https://pubmed.ncbi.nlm.nih.gov/24390935/ Hypercompact genome editor CRISPR-Casɸ By Melina Fan The ...toolbox with their discovery of CRISPR-Casɸ, a hypercompact genome editor from the Biggiephage clade of ...cells. Casɸ from Biggiephage can eliminate competing mobile genetic elements. Image by Basem Al-Shayeb...the Doudna Lab. Scientists looking for more compact genome editors due to delivery constraints and ...efficiency and reduced RNA-off target footprint compared to current N-terminal linked SaCas9 ABE variants...in screens without any reduction in performance compared to larger genome-wide libraries. Find the library...

...cells, and animals. Some of these viruses are completely inert, but others can cause diseases. Nervous...One such control is dividing up the essential components of a virus into different plasmids. For a retroviral... are not encoded in the transfer plasmid, the component that physically integrates into the host genome...genome, the virus is replication incompetent, meaning it can’t replicate and continue the infection cycle... Figure 1: Separation of retroviral packaging components onto distinct plasmids for safety.    Viral...Viral cargo-specific safety   After replication competency, the second concern with virus safety is the ...materials, you’ll need to use aerosol-tight caps, compatible with your centrifugation speed, to cover the ...

...increased versatility in probe attachment, and compatibility with far-red and near-infrared fluorophores....enable highly specific visualization of cellular components, typically a protein of interest. Fluorescent...ligands Effects on protein function and cellular components: It's important to assess how dye-conjugation... function and its propensity to bind cellular components. Some dyes target specific organelles like mitochondria...may prove challenging to stain intracellular compartments in in vivo model systems, so extra care must...Transfer (FRET) applications, the dyes need to have compatible excitation and emission profiles to enable effective...finding the right dye for your experiment more complicated. We hope this post helps you in your search for...

... a Cambrian-like explosion of software tools accompanying the CRISPR revolution. (See omictools for even... a 96-well plate and comes out of it. Biotech companies are known to have tools that do this internally...guides and primers in batch, optimized for HDR. Compared to wealth of resources for guide design, there...batch analysis because of its ease-of-use and comprehensive reports. With CrispyCrunch, you get the all ... ranking formula that does just this: Rank = (compressed CFD score) * (gaussian normalized cut-to-insert...is within the coding region, the algorithm then compares the guide sequence to every possible 23bp sequence...Francisco. Before starting his career in software, he completed a MSc in Psychology at McMaster University in ...

... stranded DNA breaks that commonly result in a complete loss of function while nuclease-deactivated Cas9...have several limitations. CRISPRko results in a complete loss of function meaning that if an essential ... more hits than the SAM library approach in a comparison study screening for vemurafenib resistance in...(Konermann et al., 2015). Root and Doench also compared the Calabrese library to screens performed with...that ORF and CRISPRa libraries can be used in complement to study gene function. Find the Calabrese pooled...gene function while using fewer sgRNAs. These new compact libraries will allow scientists to conduct genome-wide...transcriptional activation by an engineered CRISPR-Cas9 complex." Nature517.7536 (2015): 583. PubMed PMID: 25494202...

...neuronal function, with far fewer side effects compared to lesioning methods -  they may be the key to...camera or tracking software. Locomotion is a highly complex process on the level of individual neurons and ...different motivation (and brain areas associated) compared to an animal running to escape a threat. Locomotion...plus maze, the light-dark box is a simple device comprised of a brightly lit (risky) and a dark (protected... control group preferred staying in the dark compartment (Howerton at al. 2014). Addgene has several recombinase-expressing... passive mode, the stronger is the depressive component of the animal behavior. With the availability ...behavioral neuroscience to reveal insights into the complex relationship between genes, neurons, and behavior...

...mutations in the genome, but they’re inefficient compared to CRISPR/Cas9 edits that rely on double strand...only occur through editing by a APOBEC-Cas9n-UGI complex. So how exactly does this work? Figure 1:...editing efficiency can be easily determined by comparing cells expressing only eGFP expression to cells...DNA editing efficiency of APOBEC-Cas9 editisome complexes in real-time (St. Martin et al., 2019). To create...DNA editing efficiency of APOBEC-Cas9 editisome complexes. The eGFP reporters rely on the correction of ...frequency of genomic base pair edits of interest compared to cells segregated solely by a reporter of transfection... single base editing by APOBEC-Cas9 editosome complexes. Scientific Reports 9: . https://doi.org/10.1038...

...Construction of solid matrix-antibody-antigen complexes containing simian immunodeficiency virus p27 using...structures of CRBNmidi alone (A) and in a ternary complex with the multiple myeloma drug mezigdomide and ...on its own, with drug ligands, and in ternary complexes with other proteins, establishing CRBNmidi as ...matured considerably faster, with expression levels comparable to GFP during early embryogenesis (Figure 3).... through the middle of each embryo are shown, comparing GFP, wrmScarlet (mScarlet), and the new mScarlet3...Merritt D.M., Pe K., Cesar M., Hobert O. (2024) Comparative analysis of new mScarlet-based red fluorescent...than FRET-based mechanosensors (~60 kDa) and are compatible with common microscopy methods. Plus, the luminescence...

... newly formed compensation committee. The compensation committee creates a compensation philosophy that...employee compensation and also serves as an objective body to oversee some of Addgene’s compensation decisions... environment.   What do you hope Addgene will accomplish in the near future? Right now it’s a matter of...

...the N2 gene of SARS-CoV-2. If the Cas12a-sgRNA complex binds to the target sequences, Cas12a activity ... amplified region of interest. Cas13 uses a complementary crRNA sequence to bind to the sample and upon...steps and the Cas protein used. Here, all the components needed for the reaction are mixed in a single...incubated at one temperature. Two Cas12a-crRNA complexes are designed, each binding to one strand of the... RPA primers, the Cas12a-crRNA binds to the complementary region in the target sequence and this activity... substrates for Cas13. Cas13, upon finding complementarity between the gRNA and a given fluorescently ...streptavidin coated beads. Fluorescently labelled dCas9 complexes containing sgRNA that have no mismatches bind ...

...websites compiling antibody data that can help you compare antibodies from different companies, such as... a fluorophore (Figure 1).   Figure 1: Comparison between how direct immunofluorescence and indirect....  How do you find validated antibodies? Many companies that produce and distribute antibodies validate...through their publications or as a review on a company website. It is important to review the data and... a couple of antibodies that seem promising to compare in your own lab. Even with all of the resources...

...tough. Is the content accurate? Are we conveying complex ideas in the easiest way possible? Are the shots...Jennifer. I had finally found my footing with the company. I’d identified the ways to get the most out the...things like lighting, camera angle, and shot composition.  So back to the drawing board.  Cycle 3: A Return... and availability, a traditional solution was completely out of reach for Addgene.  So, onto research!...  Figure 4: The remote directing setup, complete with wireless video receiver, camera control, ...Future! Figure 5: The mobile monitor and teleprompter being repurposed for a different shoot to create...monitor stand into every shoot. I also added a teleprompter to the mix, adapting the setup to run some remote...

...lengths to keep accurate records, but given the complexities of modern research, it’s often not enough. A... have software tools that help us efficiently accomplish our daily goals. These tools remind us of our...others using excel sheets or organizing folders in complex ways, there are solutions that bring more to the...-based solutions or open source solutions. In comparison to paper-based record keeping, ELNs offer functionalities... if you are working for a large institution or company, site licenses might be available as well. In most...information. Does your lab need to meet any specific compliance requirements – e.g. 21 CFR part 11, HIPAA? Answering...of time and effort and give the lab a serious competitive advantage.  Many thanks to our guest blogger...

...are designed to be complementary, but each primer may extend beyond the complementary region as long as...products (green) are digested with enzyme “A” as compared to the products of parental plasmid digestion ... Primer Design As a rule of thumb, 11 bp of complementary sequence on either side of the desired mutation...template should be isolated from a methylation competent bacterial strain (e.g. DH5α which is dam+). Methylation...A-overhangs (a feature used in TA-cloning). Such non-complementary bases at the 3’-end interfere with plasmid reconstitution...that any residual parental plasmid (usually from incomplete DpnI digest) can also form colonies under these...

...5). Optimization of this system defined three components required for translation (Figure 1): A tRNA ...the pSer-incorporated polypeptide. These three components were shown to effectively incorporate phosphoserine... the phosphoserine OTS, Sep-OTS, contains the components required for the translation of pSer-incorporated...host strain for pSer-incorporation The second component necessary for phosphoserine incorporation is a...enzyme function, protein interaction, etc. by comparison to non-phosphorylated or ‘wild type’ control ...substitution. Alternatively, you can express the completely wild type protein from the same plasmid.  Upon...dephosphorylation reaction can be tracked through the comparison of phospho:non-phospho species on PhosTag gels...