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...Fragment? Consider the classic IgG antibody (Figure 1): it is composed of four peptide chains (two identical...that actually bind antigens.      Figure 1: IgG antibody structure with important features labeled...fragments described above can be used for similar applications and have similar advantages, though with some...

...Glycine-HCl 25–200 mM SDS 0.1%–1% (w/v) Tween 20 (optional) 1% (v/v) HCl Adjust to pH 2.0... tool to identify a protein of interest in a complicated solution like cell lysate. But they can be a ...

...forms of science communication. Click here to listen to the interview with Tom McFadden!   1. Rosalind Franklin... from Addgene.      Check out more science education tools like Verse Two and Fill in the Planck at...with us. Tom McFadden is a YouTuber and science educator looking for creative ways to implement the next...next generation science standards and communicate science. Learn more about Tom on his youtube channel ...Community Check out All of Our Podcasts Find Great Educational Content in our Plasmids 101 Series Additional...

... Joachim Goedhart Podcast breakdown Intro Chapter 1 - How Joachim ended up at the University of Amsterdam...Twitter and has helped us update many of our educational resources. Listen to hear all about Professor...speeds science. Chapter 8 - The importance of communicating research findings through social media and other...

...phosphorylated DNA termini to 3 ́-OH DNA termini (1-4). In vitro, ligases (notably T4 DNA ligase) are ... enzymology of DNA repair enzymes.     References 1. Shuman, Stewart, and Christopher D. Lima. "The polynucleotide...thermostable DNA ligase-encoding gene." Gene 109.1 (1991): 1-11. PubMed PMID: 1756968. 7. Lauer, G., et al. "Cloning...or DNA base pair mismatches (8-11). For some applications, there cannot be any end-joining activity, and...regions. Why use a high fidelity ligase? Numerous applications have been developed to take advantage of the...Taq and other thermostable DNA ligases. These applications detect specific nucleotide sequences with high...rounds of melting and annealing allows linear amplification of the probe ligation product. With the right...

...found climbing mountains or playing with his two cats. Keira Havens grew up in Hawaii where she was fascinated...challenges involved in plant synthetic biology. Challenge #1: Public perception of synthetic biology One of the...biotechnologists face is public perception of genetic modification (GM) and synthetic biology. The mistrust and...cannot build what we do not understand, and the complicated and unique metabolic processes of plants are ...

...CRISPR tools.  Distributing and depositing In Figure 1, you can see cumulative numbers of CRISPR plasmid ...new CRISPR depositors each year.    Figure 1: Addgene CRISPR distribution and deposits. (A) Cumulative...estimated scatterplot smoothing. (C) Number of requests per year for pooled libraries, categorized by function...insertion via a chemical modifier. We used these categories to see what kinds of CRISPR tools people were... pooled libraries, although we used different categories (Figure 2C). Note that in Figure 2, the numbers...Cumulative number of deposited CRISPR items, categorized by CRISPR function, by year. (B) Number of requests... years.  Libraries are divided into different categories based on function: knockout (KO), activate, inhibit...

...interrogated by the optical system (Figure 1).   Figure 1: Cells expressing a particular surface ...populations of cells in a complex tissue or sample (1). When combined with cell sorters, this technology... it passes through and scatters in different directions. The degree of scatter is captured by detectors...Light that scatters sideways to the path that it was initially traveling is called side scatter (SSC) and...granulocytes in a plot of forward scatter (FSC) versus side scatter (SSC). Users can gate around their...protein composition. It is used for a variety of applications such as measuring target protein expression ...expression levels, assessing post-translational modifications, determining cell health, analyzing cell cycle stages...

...editors (ABE) mediate A•T-to-G•C base changes (Figure 1), but it can be challenging to make these base changes...dCas to the genomic DNA of interest  Figure 1. When ABEs are in action they bind DNA so that the... from E.coli, named TadA7.10 in ABE7.10)  a catalytically impaired Cas protein (dCas or Cas nickase)  ...accidental insertions and deletions (indel) by the catalytically impaired D10A nickase from ABE7.10 can be a ...the authors created ABE8 constructs using a catalytically “dead” S. pyogenes Cas9 (dC9-ABE8.x-m/d). The...these variants, ABE8 could target many additional locations in the genome. Compared to ABE7.10, NG-ABE8.xm... reduced off-target editing frequencies. For applications that require high DNA editing specificity, for...

...fluorophores are constantly being engineered. Table 1 lists a few common fluorescent proteins and their ...definitions found in Table 2 may not apply. Table 1: The specific mutations comprising common fluorophores...transgenic mice. Purification: GFP can be used as a general epitope tag for protein purification and a number...their experimental needs, and some of the common applications in the lab. Why Green Fluorescent Protein?...to maintain fluorescent activity; very little truncation is tolerated, however, point mutations are acceptable...Re-engineering GFP to increase its range of color and application Almost as soon as its sequence was elucidated...others) Neutral mutations    A multitude of applications Due to its size and ease of use, GFP and other...

...cleavage (Harrington et al., 2017). Figure 1: Anti-CRISPR proteins (Acr's) can block CRISPR activity... ways to anti-CRISPR proteins in your experiments 1. Reduce off-target effects Prolonged Cas activity ..., 2018). For example, AcrIIA4 mentioned above indicates that it’s the fourth Acr protein discovered that...viral particles that have the gene deletion could replicate when challenged with the S. islandicus CRISPR-...B, Bondy-Denomy J (2020) Anti-CRISPR protein applications: natural brakes for CRISPR-Cas technologies....light-inducible Cas9 activity? Check out these these plasmids Catch Up on Your CRISPR Background with Our Guide Pages...

...crevices proteins on the cell surface. Figure 1: Comparison of common antibody, HCab, and nanobody...Humana Press, pp 133–148. https://doi.org/10.1007/978-1-62703-293-3_10 Walter J et al. (2020) Synthetic nanobodies...Starting with libraries and a biotinylated (for purification purposes) protein target of interest, the protocol...-30) of sybody candidates are sequenced for identification. All unique sybodies can be subcloned into ...2020) Generation of synthetic nanobodies against delicate proteins. Nature Protocols 15:1707–1741. https...Light to Manipulate Cell Signaling and Protein Purification RANbodies: Reporter Nanobody Fusions Plasmids...

...used in way more applications.   Fig. 1 Making a Protein Tag (Abridged) (1) Locate a promising monoclonal... by its antibody without additional chemical modification steps and when the epitope was within the tagged...

...Updated Jun 1, 2021 by Meghan Rego. Reproducible data are key to science, so scientists are used to repeating...the DNA packaged inside the viral particles (Figure 1). Briefly, packaged DNA is extracted from purified...These impurities cannot be removed during the purification step because they are inside the virus itself...very low concentration. So how do we make sure to catch all of these potential mishaps before distributing...Guerin et al., 2020).  In addition to genome identification, we developed custom open-access Python scripts...

...Podcast breakdown Intro Chapter 1: Who is Crystal Shih? Crystal's educational background and position at Novartis...Chapter 7: How Crystal developed her science communication skills and how they've helped her at Novartis...

...for example, split cells on a Monday diluting them 1:10, you should be able to split on the following Thursday...Immunology. You can find Sana on LinkedIn.   References 1. Yin, R., et al. "Gene expression profiling analysis...results. Most cell culture media contain the pH indicator phenol red (pH range 6.0 (yellow) - 8.0 (red))...Anything between orange and light orange usually indicates that the cells have grown enough, are making their...

...DYKDDDDK or DYKDDDK 1 Affinity and Purification Good for antibody-based purification; has inherent enterokinase...does not greatly alter the size of the tag.   Table 1: Common protein tags Tag Epitope Mass (kDa) Function...site GST Large Protein 26 Purification and Stability Good for purification with glutathione; protects...used for purification as elution requires low pH poly His HHHHHH  0.8 Affinity and Purification Very small...GKPIPNPLLGLDST   1.4 Affinity and Purification Good for antibody-based purification Combo and cleavage tags...multitude of uses including (but not limited to) purification, detection, solubilization, localization, or...fusion proteins to aid in the expression and purification of recombinant proteins. Tags for stability...

... edit.   Figure 1: Choosing the best PE6 variant for your application. Image reused form Doman...delivery by modular envelope design. Nature Communications, 14(1), 5141. https://doi.org/10.1038/s41467-023... CreVLPs, and eVLPs for in vivo and in vitro applications.   Figure 2: Modular features of DIRECTED...approach for experimental therapies and medical applications.  Find DIRECTED plasmids here! Strebinger, D...interactions or map a local proteome, but some applications have been challenging or inaccessible. The Ting... (which had therefore been trafficked between locations). Figure 3: A) LOV-Turbo becomes active...active when illuminated with blue light, catalyzing the biotinylation of nearby proteins. B) TransitID uses...

...membrane targetin sequence derived either from p63(1-29) and Lck. Image from Chertkova et al., 2020. ...Reports. 2020. https://doi.org/10.1038/s41598-020-57500-1 The CRISPR corner By Jennifer Tsang Here are a few...Fluorescent tools for mammalian cells Nanobody purification CRISPRi and CRISPRa system for hPSCs The CRISPR...CRISPR corner New from the viral service Faster catalysis after yeast-based protein evolution By Joanne ...concept, Sanchez and Ting sought to improve the catalytic efficiency of tobacco etch virus protease (TEV...activity with a fluorescent protein--no affinity purification required! The team used other molecular techniques...evolution platform to generate TEV protease variants. Catalysis by TEV protease allows a transcription factor ...

...Aptamers and Their Biological Applications.” Sensors (Basel). 2012; 12(1): 612–631. PubMed PMID: 22368488...be selected for these small ligands. References 1. Ma, Hanhui, et al. “Multicolor CRISPR labeling of... Aptamer biosensors are used in a variety of applications, including detection of environmental pollutants... opens aptamers for use in many entirely new applications including aptamer based fluorescent biosensors... render this approach less sensitive in most applications. Learn About Fluorescent Biosensors Light-up..., MG proved to be too toxic for many in vivo applications, but this work proved the concept that RNA could...and biochips. Ultimately, there are just some applications that lend themselves to aptamer-based instead...