...surprising discovery that both purified CPP-tagged protein and an untagged control could enter cultured cells ...derived from the same cell type; however, their usage has been limited by technical problems. For instance,...binary systems in which transient protein expression has a long-lasting effect, such as Cre- or Cas9-mediated...effects. Third, protein purification (once a protocol has been established) may be less laborious than viral...used for protein purification must be helping the untagged protein enter cells. Subsequent work found that...stem cells, they found very high recombination percentages (78-79% after two rounds of transduction.) Compared...

...cells according to whether or not they have been tagged with a fluorescent protein or dye. It separates... distinguishing cell populations that have been tagged with different fluorescent proteins or dyes. 3.... cell mobility within these complex environments has never been easy and, for many years, researchers ...Intravital imaging powered by two-photon microscopy has uncovered host-pathogen interactions leading to the...proteins can be used in your experiments, experience has shown that some are more reliable and more efficient...important to finely tune the excitation wavelength as it has been shown that even a small incremental change of...this system called Brainbow, the Ubow mouse strain has been created to fate map Langerhans cells and follicular...

... be selected, since it cleaves at the stop codon TAG. Design your repair template with 55-100 base...mScarlet).   Plasmid ID Plasmid Name Tags 69090 pMJ915 MBP (N terminal); 6xHis (N ...CRISPR genome editing has made it easier to create knockout alleles in a variety of species, including...including creating Cre-dependent conditional alleles, has remained a challenge. Enter Easi-CRISPR: a method...left homology arm guanine (i.e. GG); this design has been shown to increase T7 promoter activity. To avoid...

...into a vector with a large tag at the N-terminus, but you don’t have the tag already included in the vector...you can insert both the gene of interest and the tag sequences into the vector in one step without scars...where the vector that you want to use is very large, has a high GC content, contains a lot of repeats--any...In this blog post, I will go over some advantages, disadvantages, and examples of how scientists are using... common, and for good reason. They offer many advantages over the traditional restriction enzyme cloning...thereof) nor a serious time commitment. Another advantage is that this process makes it easy to generate...will be using. Some of these products may offer advantages that Gibson assembly does not such as increased...

...respectively. mNeonGreen can be used as a fusion tag for a wide range of applications from traditional...single-molecule super resolution imaging. mNeonGreen also has little sequence identity to other common fluorescent...fold brighter than EGFP and is photostable. It also has narrow excitation and emission peaks (max excitation...was then screened through successive rounds of mutagenesis for the brightest teal fluorescent proteins that...scientists from Florida State University, they used a mutagenesis approach and found that specific amino acid changes...

...artifacts, it has better folding and maturation kinetics than other dark fluorescent proteins, it has good signal...encoding alternative promoters and/or fluorescent tags are available at Addgene. Shemesh OA, et. al. ...Technology (KAIST) and Institute for Basic Science (IBS) has developed another useful optogenetic tool that enables...facilitate future work with this goal, the Barrick lab has deposited the pDCAF3-IS5 plasmid containing a synthetic...operon originates from Pseudomonas putida CBB5 and has been tailored for expression in E. coli. This operon... contributed by Christina Mork The Murakoshi lab has developed two new FLIM-FRET acceptors, ShadowG, a... due to its very low quantum efficiency. ShadowY has a lower quantum yield and higher absorption than ...

...used for PL were APEX2 and BioID. APEX2 is fast, tagging proximal proteins within minutes, but requires ...being targeted via RNAi. Recently the Vellai lab has created an improved RNAi vector that leads to significantly...sequence of the toehold switch. For example, Green has also developed toehold switches to detect Zika virus...protein motifs than previously thought. Sidhu’s group has made several hundred SH3 domain containing plasmids.... In addition to these constructs, the Sidhu lab has made hundreds of additional plasmids available through...infect millions annually. Because it is highly contagious, it is essential to contain outbreaks when they...

... expressing fluorescent proteins will be used to tag individual strains of bacteria. Their interactions... S. University of Baroda, India  Devarshi Gajjar has 10 years experience in the field of bacterial and...biomedicine and astrobiology. During the last 11 years he has studied various aspects extreme life on earth including...educational resources for scientists globally, Addgene has partnered with various community organizations and...

...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...

...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...

...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...

...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...

...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...

...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...

...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...