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Showing: 601 - 640 of 1100 results

  1. An Addgene Co-op: The Intersection of Fun, Friendship, and Work

    Type
    Blog Post
    ...but overwhelmed; I had no idea where to start. Thankfully, my managers set aside time for me to complete...the lab! I got to assist production on a protocol video. Don't worry - no pipettes were harmed in the making...allowed me to succeed. As a team, we talked through slide design, discussion items, big takeaways, and timing...depositor welcome kits, note taking for protocol video production, and keeping inventory of our merchandise...talk to. This six month experience made me more confident in myself, my abilities, and my future - something...

  2. Five Questions to Ask If You're Deciding Between a Master's or PhD Program

    Type
    Blog Post
    ... at the Addgene Science Career Guide eBook Addgene Science Career Guide eBook.   Resources Resources... a master? Here are a few questions to help you decide if a master’s might be for you. 1. Are you interested...path one you could see yourself taking? If so, consider setting up an informational interview with them...your interests and career goals. You might still decide to earn a PhD, but if I had to do grad school over...eBooks from Addgene, including the Science Career Guide\  ...

  3. The Stingy Scientist: How the Baby Gel Box Was Born

    Type
    Blog Post
    ...pour over the sides.  These tiny gels and tiny lanes required less agar, less ethidium bromide, less stop...discards according to the Cambridge, Massachusetts guidelines and wear an extra sweater so we don’t have to...smaller gel apparatus until I settled on using glass slides. With just the right amount of agarose, I could...could pour a very thin gel on a large glass slide using only surface tension.  It's a bit tricky to pour ...plastic and solder this thing together.  One of the downsides—there was no cover so you had to be careful not...

  4. Bright Monomeric Fluorescent Proteins: mNeonGreen, mTFP1, and mWasabi

    Type
    Blog Post
    ...in 2012. Using structural modeling and structure-guided direct evolution, Nathan Shaner and colleagues ...respectively. mNeonGreen can be used as a fusion tag for a wide range of applications from traditional imaging to...resolution imaging. mNeonGreen also has little sequence identity to other common fluorescent proteins and therefore...The team that developed mTFP1 further sought to identify why this protein was blue-shifted compared to ...blue-shifting more than others. During this process, they identified mWasabi, a bright, monomeric green fluorescent...variants of Clavularia cyan fluorescent protein: identification of the molecular determinants of color and ...

  5. Natural or synthetic? How Addgene’s dataset reveals trends in biological innovation

    Type
    Blog Post
    ... sequence identity was a different story - the team found that the percent sequence identity between the...) and comparative properties (percent sequence identity and percent query coverage) that could be determined...synthetic or natural. They saw that a percent sequence identity below 85% correlates with synthetic sequences.... The researchers then used their classifier to identify synthetic and natural sequences in Addgene’s database...have practical applications in monitoring the accidental or intentional release of engineered organisms...

  6. Addgene's Top 10 Blog Posts of 2015

    Type
    Blog Post
    ...per month in November 2015. We will continue to provide you with more fascinating, informative, and actionable...technology in their own labs. We’re overjoyed to provide you with these fantastic educational resources ... you’d like to see on the blog or to contribute ideas for your own guest posts by e-mailing us at blog...inversions, deletions, and translocations. It also provides researchers with many different ways to control...the right one. In this post, Addgenie Melina Fan guides us through the basics of the validation process...experiment? In this post, Addgenie Chari Cortez guides us through some common controls and why you might...

  7. Creating accessible biology activities in schools with BioBits

    Type
    Blog Post
    ...enough to have the funding and the resources to provide them to their students. My desire to create a set... cell-free (FD-CF) components. Making proteins outside of a cell - cell free extracts Typically, if you...used in high school classrooms, but it could be widely adapted and used in a variety of settings. If you...are an instructor at the college level, perhaps consider using some elements of BioBits to transition your...researcher interested in adapting your research ideas into hands-on educational activities using the BioBits... Resources on Addgene.org Browse our protocol videos to learn simple lab techniques Check out our molecular...

  8. Plasmids 101: Gram Negative and Positive Bacteria

    Type
    Blog Post
    ...stain for 1 minute. Rinse the slide briefly with tap water. Flood the slide with Gram's iodine solution....staphylococci and streptococci. Gram negative bacteria, or diderms, have a thinner peptidoglycan membrane and an outer...safranin counterstain for 30 - 60 seconds. Rinse the slide in tap water until the runoff is clear, blot gently...types. In water, the dye dissociates into CV+ and Chloride (Cl-) ions. When the negatively charged iodine...into a specific bacterium, it can be important to consider the type of bacteria it is being used in, and ...

  9. 5 Great Apps for Lab Life

    Type
    Blog Post
    ...interwebs for lab resources, and instead conveniently provide them on your smart phone. 1. Lab Archives by LabArchives...Gibco Cell culture manual and basic instructional videos. The search function in the manual just highlights...with NEB enzymes, the app is quite helpful and provides a list of directions including which buffers to...primer, you only need to skim the alphabet on the side of the screen to find the sequence for your enzyme...explain the properties of or draw an amino acid or peptide for some reason. It contains all twenty amino acids...allow you to write your colleagues’ names in a polypeptide chain, except if their names have a B, J, X, ...

  10. R Bodies: Membrane-Rupturing Microscopic Tools

    Type
    Blog Post
    ... proteins. Video by ACS. For a more cartoony explanation of R-bodies, check out this video. Most types...by propagating through an enormous lattice of identical subunits. A brief history of R bodies R bodies...proteins and remain functional, 2) break membranes outside of their natural context or 3) be modified to extend...fluorescent protein fusions allowed us to image R bodies inside cells, and these constructs may be useful to future...soluble when extended (see figure to the right), we identified R-body mutants that extend at both lower and ...

  11. Management for Scientists: Delegating is Key

    Type
    Blog Post
    ... More people in leadership roles leads to more ideas, initiative and creative thinking. The team members... must usually take on all tasks that involve confidential information. No matter who you delegate to, ...can help ensure successful results.  Can you provide the information and resources needed for the assignment...Setting someone up for failure by delegating is a bad idea. How do I make sure it goes well? As with most ...even if it is just 15 minutes a week. Clearly identify constraints and boundaries. Where are the lines...issues do you want to know about right away? Provide adequate support. The manager must be available...

  12. Finding and Joining Your Dream Lab

    Type
    Blog Post
    ... perhaps surprising, yet important, factors to consider when seeking your dream lab. The principal investigator's...benefits. Therefore, lab selection should focus on identifying a lab that fits both work style and development...development needs. These two factors will provide a solid foundation on which to thrive during time spent as ... factor. The following questions can serve as a guide for the quest of a perfect "fit". How competitive...want a non-academic career path?  If you are considering this, make sure to choose an advisor that supports... into place frequently and you sometimes feel confident in your abilities. You have intense periods of...

  13. From Lab Bench to Leadership: Transitioning into Management as a PhD

    Type
    Blog Post
    ...arises when managers fail to set expectations or provide timely feedback, leading to unproductive dynamics... mentors who have made similar transitions can provide you with insights, lessons from their experiences...work. On the other hand, it’s perfectly valid to decide that managing isn’t for you at this point in your...focusing on molecular biology and metabolism. He now guides fellow PhDs in their careers with actionable steps...Coursera, LinkedIn Learning, or edX. Podcasts: HBR IdeaCast Coaching for Leaders More resources on the ...Makes a Good Leader Anyways?  Transferable Skills Guide: Managing a Team  ...

  14. GCE4All: Making Genetic Code Expansion Accessible

    Type
    Blog Post
    ...impact both translation and fidelity of translation, leading to system-wide effects. And on top of that...modification, or add in fluorescent or chemically reactive side chains, all of which can help determine structure...reagents accessible to your lab. The solution GCE provides almost endless possibilities for researchers, ...researchers can easily implement in their own labs. They provide resources, knowledge, and most importantly, validated...that is by using Addgene’s Blue Flame award to identify highly requested GCE plasmids. But they’d also...

  15. Hot Plasmids December 2018 - C. elegans RNAi, TurboID, Detecting Viral Outbreaks, and Protein Binding Specificity Tools

    Type
    Blog Post
    ...through our hot plasmids articles. These articles provide brief summaries of recent plasmid deposits and ...to prevent the incorporation of non-specific nucleotides into the dsRNA leading to increased RNAi efficiency...This tetramer cleaves chlorophenol red-b-D-galactopyranoside on the assay paper turning it purple, allowing...30370338.  SH3 domain collection reveals novel peptide ligand motifs Article contributed by Michael Lemieux...specificity landscape of the SH3 proteins using peptide-phage display and deep sequencing, revealing that...protein stability, small molecule binding screens, peptide arrays, and phage display selections. In addition...

  16. Top Requested Lentivirus and AAV of 2016

    Type
    Blog Post
    ...With this initial success, we will continue to provide and expand this diverse and useful collection of...inventory is based on CRISPR tools, though we do provide a few non-CRISPR related viral preps. Cas9 is the...with a second vector that delivers gRNA, such as lentiGuide-Puro. The alternative single-vector systems typically...gRNA lentiviruses easily made in your own lab. We provide protocols and useful tips for generating your own...exclusively activated by the small molecule clozapine N-oxide (CNO) (Krashes et al., 2011). Viruses expressing..., Shalem O, Zhang F. Improved vectors and genome-wide libraries for CRISPR screening. Nat Methods. 2014...

  17. 3 Challenges in Plant Synthetic Biology

    Type
    Blog Post
    ...biotechnology that are ripe with opportunity. We’ve decided to jump into one of those unexplored areas with...food, not medicine, and not mandatory – people can decide for themselves if they want this beautiful flower...begun to pivot towards plant biotechnology and provide a much needed boost to basic molecular plant research...are well understood. Additional work has also identified and characterized mechanisms and components of...you to our guest bloggers! Nikolai Braun grew up outside of Washington D.C. and has always enjoyed exploring...laboratory and received her M.S. from CSU in 2014. Outside of the lab, Keira enjoys spending time with her...

  18. Plasmids 101: Modular Cloning Applications and Kits

    Type
    Blog Post
    ...construct. Once you’ve identified your experimental requirements, it’s a good idea to select an available...biofuels, and natural products without the undesirable side products generated during chemical synthesis of ...find the perfect combination. The EcoFlex Kit has a wide array of DNA parts that can be assembled into individual... are endless: genetic circuits with multiplexed guides, mini screening libraries, direct edit optimization...cells. Need more MoClo? Check out Addgene's MoClo Guide! Customizing MoClo for your needs As you probably...Resources on Addgene.org Modular Cloning (MoClo) Guide Addgene’s eBook Collection More resources on the...

  19. Special Delivery: Fluorophore Targeting for FRET Studies

    Type
    Blog Post
    ... donor and acceptor fluorophores, it’s best to consider these cases separately when talking about the ...the tools that are available. Let’s consider the donor first where labeling specificity is most important...hardwire the molecule directly into the protein’s peptide sequence. Thus, all synthesized target protein ...addition, labeling contrast can be improved using disulfide-containing compounds such as British anti-Lewisite...described above can be paired together to measure a wide range of molecular distances (see Table). However...covalent labeling of recombinant protein molecules inside live cells." Science281.5374 (1998): 269-272. PubMed...Protein Fusions Check out Our Fluorescent Protein Guide ...

  20. dTAG - You're it!

    Type
    Blog Post
    ...degradation. Remarkably, small molecule degraders provide distinct advantages over pharmacological inhibitors...Nabet et al., Nature Chemical Biology additionally provides detailed information on cloning procedures used...molecules by conjugating a CRBN-binding ligand (thalidomide) to an FKBP12F36V binding ligand (ortho-AP1867...commercially available in the near future, and we will provide an update with vendor and catalog numbers. As we...or Jay Bradner ([email protected]). Considerations when getting the dTAG system up and running...while quantitative proteomics experiments will provide the best confirmation of selective FKBP12F36V fusion...

  21. Antibodies 101: Secondary Antibodies

    Type
    Blog Post
    ...signaling molecules to identify each protein of interest, this gel-based method identifies proteins solely by... blog posts, then you likely already have a good idea of the basics of immunoimaging. A scientist conjugates...place over multiple days. Increased run time is considered one of the major drawbacks of the indirect method...interest - the secondary antibody needs careful consideration before selection. If all the primary antibodies...themselves able to approach immunoimaging assays more confidently, collect a broader array of data, and occasionally...

  22. What's New in CRISPR - November 2021

    Type
    Blog Post
    ...recently identified Cas12f systems from uncultivated archaea are much smaller and would be ideal for such...different labs have recently published work that provides a solution to this problem. A miniature CRISPR-Cas...editing with a hypercompact Cas12f1 and engineered guide RNAs Yong-Sam Kim’s lab also created a miniature...Biotechnology paper showcases an improved design of the guide RNA that caps the 3’ end with a structured RNA motif...sheet Resources on Addgene.org Read our CRISPR guide Find CRISPR plasmids by function Find recently deposited...

  23. All in a Twist: dsRNA

    Type
    Blog Post
    ...into agriculture as species-specific pesticides and insecticides, an advancement over previous chemical...which detect dsRNA in the cytoplasm and use it as a guide to complementary RNA sequence. Once a compatible...processes and functions, but they also have a darker side. When ssRNA engages one strand of a dsDNA molecule...if endogenously synthesized dsRNA finds its way outside of the mitochondria, the same inflammatory response...or anticipating this response are important considerations when using these reagents. RNA:DNA hybrids...

  24. CRISPR Meets Synthetic Biology: A Conversation with MIT’s Christopher Voigt

    Type
    Blog Post
    ...assembling the toolbox needed for anyone to begin considering genetic engineering projects in a very big way...limitless number of DNA binding proteins with Cas9 guided by different promoters. The challenge in building...therapeutics, if you want to program a cell to identify and target a malignant cell, then you’d need the.... You couldn’t do it without repeating the same guide RNA twice. The second problem was the shape of the...also difficult to build larger circuits with many guide RNAs connected because the signal degrades at each...now? Voigt: We are working on this now, we have ideas, but so far nothing that has worked. I don’t have...

  25. Engaging with science and society at pgEd

    Type
    Blog Post
    ... religious affiliation, and ethnic or personal identity. We do this through a variety of activities, including...experiment, the case of Henrietta Lacks, the Havasupai incident, and the sterilization of women in marginalized...with different organizations and individuals, to identify ways of collaborating, and to organize events ...Having a broad interest and awareness of issues outside of your field of research will allow you to better...better engage these diverse audiences and to identify issues or topics that are matters of concern for ... future work in these areas, and in helping you decide whether you enjoy this kind of work. It is also...

  26. Educational Planarian Tools for Teaching Developmental Biology Techniques

    Type
    Blog Post
    ... creatures commonly found in freshwater ponds, provide a good potential solution to this problem. Planarians...companion resource called “cutting class” which provides more detailed materials and protocol resources...synthetically made RNA (riboprobes) with modified nucleotides (for example, Digoxigenin-labelled Uracil) that...dsRNA feedings. Some of the phenotypes are not evident until after amputation. For example, the dramatic...odf2 gene knockdown results in an observable “sidewinder” phenotype that disrupts wildtype locomotion....dsRNAs into PR-T4P. The Sánchez lab protocol also provides step-by-step instructions for the “the bravest...

  27. Hot Plasmids May 2018 - Optogenetics, Decaffeination, Biosensors, and Fluorescent Protein Tools

    Type
    Blog Post
    ...through our hot plasmids articles. These articles provide brief summaries of recent plasmid deposits and ..., CRY2clust, was created by adding a 9-residue peptide to the C-terminus of human codon-optimized  cryptochrome...conditions. With its additional 9 amino acid peptide, the CRY2clust domain enables fast and reversible...original version containing CRY2 without the extra peptide. CRY2clust should prove a useful tool to further...supplied in the media, these bacteria are unable to divide. However, if fueled with caffeine and equipped ...Dronpa flanked a PKA substrate fused to the phosphopeptide binding domain FHA. The FHA1 domain binds to...

  28. Corralling and Cornering the Goldilocks Antibody

    Type
    Blog Post
    ...scientists’ pockets. They allow researchers to identify and isolate very specific biological targets. ...can then be used by thousands of scientists to confidently choose reliable and consistent antibodies for...antibodies, creating a potentially unlimited supply of identical antibodies (See Fig 1.). rAbs are a relatively...collaborates with these companies to create and provide general availability of their recombinant antibodies...creating a database for goldilocks antibodies and, alongside that, a publicly available 2D/3D mouse atlas from...atlas will enable scientists to have an intuitive guide for checking their staining and neuroanatomy without...

  29. CRISPR 101: Targeting Non-Coding RNAs with CRISPR/Cas9

    Type
    Blog Post
    ...that are around 20 base pairs long. Using a single guide RNA (sgRNA) to create a double-strand break works...non-coding RNAs (lncRNA). lncRNAs are over 200 nucleotides in length and lack an open reading frame, so ... a number of different options. By carefully considering your ncRNA of interest, you should be able to...references Resources on addgene.org Addgene's CRISPR Guide Addgene's CRISPR 101 eBook CRISPR resources More...Doudna JA, Charpentier E. A Programmable Dual-RNA–Guided DNA Endonuclease in Adaptive Bacterial Immunity...051410-092902 Liu Y, Cao Z, Wang Y, et al. Genome-wide screening for functional long noncoding RNAs in ...

  30. Plasmids 101: Modular Cloning

    Type
    Blog Post
    ...given peptide can easily be tested by generating Level 1 TUs with unique promoters, and the ideal phosphatase...behave in a cell. For unpredictable cases, MoClo provides two answers: first, sets of parts with characterized...for more complex CRISPR experiments (e.g., many guides/parts delivered) in certain model organisms. In...combinations of different candidate genes at once to identify the best cocktail of genes when the relationship...Resources on Addgene.org Addgene’s Modular Cloning Guide EcoFlex MoClo Kit CIDAR MoClo Kit MoClo Plant Parts...

  31. Plasmids 101: FLEx Vectors

    Type
    Blog Post
    ...using either loxP or lox511 sites, leaving two identical sites on one end of the DNA fragment (Figure 1...recombination event then excises the DNA between the identical loxP or lox511 sites (Figure 1, bottom). Since...only one loxP and lox511 site will remain on either side of the DNA fragment, any additional recombination...genetics, FLEx is currently being used to study a wide variety of in vivo conditional gene rescue experiments... silent until a cell or tissue-specific Cre is provided. Since the FLEx switch ensures Cre-dependent reporter...inverse Orientation), FLEx systems are generally considered "Cre-On" meaning that your gene of interest starts...

  32. Illuminating Epigenetics with A FRET Based Biosensor

    Type
    Blog Post
    ... of the DNA with a high frequency of C and G nucleotides). The conversion of cytosine to 5-methylcytosine... to study epigenetics Epigenetic research uses a wide range of molecular biologic techniques to further...enzyme digestion, DNA adenine methyltransferase identification, and bisulfite sequencing. However none of ...consists of one substrate (the histone-derived peptide) which can localise the construct specifically ..., which binds selectively to lysine-methylated peptides. This construct is in the middle of a FRET pair...When methylation occurs at the histone-derived peptide, the methyllysine binding domain causes a structural...

  33. Multiplexed Capture of Promoter-enhancer 3D Chromatin Structures Using CRISPR

    Type
    Blog Post
    ...While “pictures” generated with current 3C methods provide useful information about chromatin interactions...streptavidin IP. Biotinylated dCas9 goes along for the ride throughout all of these steps, until its connection...off-target sites. Long-range chromatin interactions identified with the multiplexed CAPTURE 2.0 largely replicated...replicated interactions previously identified with the original CAPTURE method. Both CAPTURE methods outperformed...both the number of unique chromatin interactions identified and the level of on-target enrichment. 2. Determining...on Your CRISPR Background with Addgene’s CRISPR Guide Page ...

  34. Chromoproteins: Colorful Proteins For Molecular Biology Experiments

    Type
    Blog Post
    ...fluorescent proteins and luminescence have been widely used in biological research for a long time, chromoproteins... give colors in ambient light. In contrast, the widely used green fluorescent protein (GFP) gives a bright...early 2000s, a handful of chromoproteins were identified in sea anemone (Chan et al., 2006). Why do these... sequencing are more definitive, but visual identification is a good place to start before taking on more...could get around this problem. There is no single “ideal” chromoprotein plasmid that combines the most desirable...is an orange light-absorbing chromoprotein that Hideji Murakoshi’s lab developed as an acceptor for fluorescence...

  35. Plasmids 101: Knockout/Knock-In Plasmids

    Type
    Blog Post
    ...breaks, in which nucleotide sequences are exchanged between two similar or identical molecules of DNA....homology to the locus of interest. To give you an idea of the process, we’ll walk through an experiment... gene have been cloned into the vector on either side of an antibiotic resistance gene. To avoid selecting...HSV thymidine kinase (HSV-tk) is included just outside one of the homology arms. When we select cells ...creating knockout animals or cell lines, please consider sharing them by depositing with Addgene! Figures...

  36. Don’t FRET: Bimolecular Fluorescence Complementation Makes Visualizing Protein-Protein Interactions Easy

    Type
    Blog Post
    ...validated for BiFC (Kodama and Hu, 2012). Having a rough idea of how your PIPs might interact will allow you to...each PIP and empirically test which combination provides the best fluorescent signal. As for linker sequences...experiments. In either case, overexpression should be avoided because it can generate high background fluorescence...critical to the success of BiFC experiments. The ideal control is a mutated variant of one or both of your...that you empirically test a number of mutants to identify those that inhibit BiFC in your cellular system...endogenous PIPs is expressed at increasing levels alongside your BiFC constructs. If your PIPs actually interact...

  37. AAVs in Retinal Gene Therapy

    Type
    Blog Post
    ...was in its infancy, but the eye was identified early on as an ideal target. It is an easily accessible ... time advances in human genetics led to the identification of several disease-causing genes, including...However, AdV was abandoned early on due to its considerable immunogenicity and its transient expression....have broad tropism in the eye. AAV-2, the first identified serotype and the one used in Luxturna™, can efficiently...therapies is impressive: clinical trials for choroideremia, X-linked retinitis pigmentosa, achromatopsia...

  38. Deep Dive: Fixing and Permeabilizing for Immunofluorescence

    Type
    Blog Post
    ...method into six general steps and outlined the considerations to be made during each step. Now that you know...take a slightly deeper dive into those steps and provide you with more details to help you plan and optimize.... If your target is already on the extracellular side of the membrane, then this step may not be necessary...binding to any target protein that is still on the inside of the cell. However, assuming you do need to permeabilize... of fixatives or permeabilizers, but how do you decide which ones to use? Here are some tips for making... then optimizing from there is a good option.  Consider your target - if you are studying a membrane protein...

  39. AAVs CREATed for Gene Delivery to the CNS and PNS

    Type
    Blog Post
    ...an IV injection is an ideal way to deliver AAV to the PNS. It’s a happy accident that AAV.PHP.S targets...barrier - only those that cross the barrier can be identified by PCR later. The PNS presents a different problem...Cre-dependent selection yields AAV variants for widespread gene transfer to the adult brain. Nature biotechnology...Additional Resources on the Addgene Blog Important considerations when using AAVs Learn more about using AAV ...Vectors to use with your research. Visit Our AAV Guide Page for info on AAV components, common uses of ...

  40. Antibodies 101: Producing Recombinant Antibodies

    Type
    Blog Post
    ... Protocol Addgene's Video Antibody Purification Protocol Addgene's Antibody Guide References Trimmer JS...the periplasmic space to be oxidized and form disulfide bonds (Frenzel et. al., 2013). If you’re using...stained, the heavy and light chains become visible alongside your ladder and a protein standard. The density...calculated using imaging software. Any bands on the gel beside the heavy and light chains can also be measured...multiple contaminating bands or a few strong ones, consider increasing the number of washes performed during...
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