...210 plasmids: empty backbones, fluorescent protein tags, SARS-CoV-2 plasmids, etc. It is our hope that by...Good bye 2020! What a year it has been for all of us. As we eagerly await 2021, we've looked back on ...

...elements – promoters, origins of replication, protein tags, and antibiotic resistance markers (just to name...restricted based on experimental constraints. This has been used for a wide range of purposes, including...Cre-containing adenovirus (Ad-Cre) or AAV (AAV-pgk-Cre) has been used to successfully introduce Cre into cells... can be an effective alternative to Cre-lox, and has also been used in conjunction with it, allowing for...recombination at the loxP recognition sites. A key advantage of the system is that Cre acts independently of...in certain cell or tissue types, during certain stages of development, or by making the Cre inducible ...

... purified them to near homogeneity using a 6XHis tag, and compared their specific activities by measuring... characterization of a fusion protein: Histidine tag, malate dehydrogenase, and green fluorescent protein... from R1 institutions to community colleges, and has helped provide original research experiences for ...

...CRISPR/Cas9 has made it easier than ever to make precise, targeted genome modifications. Cas9 has been modified...same genetic locus via co-expression of an epitope tagged dCas9 and antibody-activator fusion proteins. The...which one of Cas9’s two critical enzymatic residues has been converted to an alanine (D10A or H840A), changing... for your experiment, ensure your sgRNA sequence has little homology to other regions of the genome and...looser PAM requirements compared to SpCas9. xCas9 has a PAM requirement of NG, GAA, or GAT, making it a...SpG variant, discovered by the Kleinstiver group has a PAM of NGN (Walton et al., 2020). Further, this...reverse transcriptase, and a nCas9. While this system has been shown to be less efficient than base editing...

...systematically offer thousands of constructs for tagged and untagged expression of SARS-CoV-2 proteins in bacteria...  While rapidly expanding our COVID-19 resources has been our focus for the past months, we’re excited...

...your experiment. For example, does the fluorescent tag need to be pH stable or withstand a particular solute...enzyme activity. Small molecule biosensors Addgene has many sensors that measure the concentration of small...cellular characteristics like pH, redox state, and voltage across membranes. These sensors generally follow...

...enormously plastic tool and has taken the scientific world by storm. While Cas9 has been most widely used to..., Qi LS, Weissman JS, Vale RD (2014) A Protein-Tagging System for Signal Amplification in Gene Expression... to create specific edits in DNA, there has also been significant work on constructing Cas9 transcriptional...CRISPR activation for genes with known functions has the potential to be used in therapeutics, as it can...changes in phenotypes. For example, CRISPR activation has been used to differentiate induced pluripotent stem...creation of innovative technologies. To date his group has develop several popular CRISPR-based tools for altering...Genome-wide Screens with CRISPR/Cas9 Learn How CRISPR Has Been Used with AAV Resources on Addgene.org Find...

...community has always emphasized the need for open science and collaboration. The field already has comprehensive...system a modified TIR-1 protein can degrade degron-tagged proteins in an auxin dependent manner. This system...first virtual organism in a computer.” This project has produced various resources for the worm community...

...screening, Single-cell Phenotypic Observation and Tagging with Light (SPOTlight) as a way to isolate individual...Cas12a. This is in part due to the fact that it only has a single active site. The RuvC active site in Casɸ...rely on multiple active sites. In addition, Casɸ has a minimal PAM requirement. For instance, for Casɸ...library here A new SaCas9 ABE variant (microABEI744) has improved on-target editing efficiency and reduced...

...dye, but has been modified with the addition of two iodine residues. Compared to MG, MG-2I has increased...development in real time.  In both cases, dL5** was tagged with mCerulean3 (mCer3) for visualization and expressed...xOtH5bs4HiI Advantages of MG-2I-dL5** over other photosensitizers One of the main advantages of MG-2I-dL5...

...and mammalian. These plasmids contain both untagged and tagged versions of SARS-CoV-2 ORFs and include plasmids...that have been in the repository for many years, has now grown into a collection of over 2,400 plasmids... based on 100 structure-guided spike designs and has improved protein yields and stability. This stabilized...virus spreads around the world, the spike protein has had many opportunities to evolve. Several variants... to accelerate research through sharing reagents has never been so important.     References and ...

...controlled setting. Read more: Plasmids 101: Protein Tags Produce proteins that glow so that scientists ...be used in research or for therapeutics Addgene has compiled various educational resources to facilitate...plasmid) and typically include a gene that is advantageous for survival, such as an antibiotic resistance...

...looked hard at existing tools. As has been written here on Addgene, there has been a Cambrian-like explosion... for the CZ Biohub project by Manuel Leonetti to tag all 22,000 protein coding genes in the human genome...high-throughput design and analysis of CRISPR experiments. It has certainly boosted our productivity in the Biohub.... tools which were optimized for HDR, except for TagIn. HDR requires the additional step of mutating guide...cut-to-insert distance for HDR. (The notable exception is TagIn––see their "summarisation score".) For CrispyCrunch...

...insert FLAG tags into the genome of the pathogenic fungus Aspergillus fumigatus, which has been difficult...sequence between the microhomologies.MMEJ has certain advantages over HDR. Some species don’t have good ...vectors. Addgene depositor Takashi Yamamoto’s lab has harnessed MMEJ to create a new method for CRISPR ...of press. However, this process accounts for a percentage of the mutations seen with TALENs and CRISPR....follows. For increased versatility, it would be advantageous to adapt PITCh to insert genes into AAVS1, the...using the nuclease Cpf1. Since Cpf1 cuts in a staggered pattern, it is thought to be ideal for HDR-independent...and Ling Lu. (2016). “Highly efficient CRISPR mutagenesis by microhomology-mediated end joining in Aspergillus...

...also the fastest test that has been developed so far.  A similar method has been optimized to detect the...Two-step-SHERLOCK has a sensitivity in the zeptomolar range while One-step-SHERLOCK has a sensitivity in...using lateral flow strips. Since the probes are tagged with streptavidin, the lateral flow strip first...State University. Since its appearance, SARS-CoV-2 has spread to almost every part of the world manifesting...full-fledged pandemic. Containing the spread of this virus has become an utmost priority for countries around the...the outbreak ultimately flattening the curve, it has become evident that widespread testing is crucial... 83 known COVID-positive samples tested, DETECTR has a positive prediction rate of 95% and a 100% negative...

...to allow in-frame deletions and introduce epitope tags and RNA transcription start sites. The lab further...packaging, but their activity in mammalian systems has been in question. Three different labs have recently...CRISPR-Cas system for mammalian use Stanley Qi’s lab has developed a miniature Cas system (CasMINI) engineered...

...until the pups are 2-4 weeks old and use the ear tag method (creating different patterns of ear holes ...Samantha Young. The use of CRISPR/Cas9 for gene editing has expanded since its adaptation for use in mammalian...biology surrounding a particular gene (although it has been quite successfully adapted for use in screening... method for validating your selected gRNA’s, and has the bonus benefit of supplying you with primers for...sturdier than the delicate RNA and therefore there has less chance of contamination with RNase leading to...products will give specific details on what exactly has been inserted or deleted. Each lab will use a slightly... et al. "Feasibility for a large scale mouse mutagenesis by injecting CRISPR/Cas plasmid into zygotes....

...FPs) offer scientists a simple yet powerful way to tag cellular proteins and investigate protein localization...fusion, etc). For example, by engineering PA-GFP tagged to a mitochondrial matrix protein (mito-PAGFP) ...Markus, and Wei‐Lih Lee. "Photoactivatable GFP tagging cassettes for protein‐tracking studies in the budding...high contrast ratio usually indicates that a PA-FP has a low level of spontaneous fluorescence in the absence...an FP-chimeric protein may be at any one of many stages of synthesis and degradation. For this reason it...PA-FPs by performing several rounds of random mutagenesis screening for enhanced RFP variants. PA-mCherry...negative” photoswitching. In 2008, scientists used mutagenesis screens to find a version of Dronpa that doesn't...

... applications (v82 has higher affinity for glutamate but slower kinetics; v857 has faster kinetics but...but lower affinity) and with a variety of tags for localization (PDGFR, GPI, and SGZ). Read more from the...iGluSnFR3 (Aggarwal et al. 2022). This new version has higher signal to noise ratio and is better localized...for two-photon microscopy. The resulting JEDI-2P has faster response kinetics and increased brightness...and deep-tissue imaging. Vladislav Verkhusha’s lab has generated a new genetically encoded calcium indicator...interest, it’s important to choose a biosensor that has a binding affinity close to the range of concentrations...physiology and metabolism under various conditions. LiLac has improved response size and reduced sensitivity to...

...epitope-targeting antibody that is conjugated to a biotin tag but not a fluorophore. In a separate step, cells ...interest. Indirect labeling is achieved through protein-tag or protein-protein interactions, like biotin-streptavidin...selecting the primary antibody host species, as it has to be different from the other antibodies in your...target marker, allowing for its detection. The advantage of this system is that streptavidin conjugates...the primary antibody (Figure 1b). As above, the advantage of this system is increased flexibility for panel...