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Showing: 241 - 260 of 1052 results

  1. Five Popular Model Organisms, Part 2

    Type
    Blog Post
    ...Justin Ng and Alyssa Cecchetelli Science is a constantly evolving and demanding field, requiring a variety...of biological research that has defined our understanding of biological processes and developments. Model...used as a model organism for over 60 years with a status as the most studied prokaryotic organism. With ...growth rate, small size, and low cost, it’s understandable why E. coli is the most popular bacterial organism...settings. One of its most attractive features is its stability and easily obtainable growing conditions. As a...thrive with or without oxygen. Additionally, its status as a Biosafety Level 1 microbe makes it a perfect... biofilms, and much more. Because of it’s BSL-1 status, E. coli can be used as a model to relate these...

  2. Harnessing Bacterial Toxins for Allelic Exchange

    Type
    Blog Post
    ...chloramphenicol-resistant strains, we have also included versions with gentamicin-resistance (amplified from...disrupt expression of a gene with an antibiotic resistance cassette. This leaves a “scar” in the chromosome... in the creation of scarless mutations, both in stand-alone systems (Jiang et al., 2013), and those which...be accomplished by selecting for an antibiotic resistance marker from the vector (abR, Figure 1). The resolution...This vector contains sacB, an chloramphenicol resistance cassette, and the R6K origin of replication to...replicated in the recipient, the only antibiotic resistant colonies isolated are those where the allelic ...conjugate that well. This is because in these circumstances, the conjugation can be so inefficient, that...

  3. Summer SciComm Series: Cognitive Load

    Type
    Blog Post
    ...scicomm bandstands: cognitive load.  Cognitive load The Wikipedia definition of cognitive load states: “cognitive...mental thought and effort your audience needs to understand what you’re communicating. While I’m sure that...of knowledge new to mankind. And you probably understand how much faster it would be for an expert in ... are asking your audience to take on, just to understand you at the most basic of levels.  Concepts This...lot of science communication is helping people understand new concepts. You hopefully are aware that this...foundational concepts (that they often don't!) to understand us, or being able to connect one concept to another...for your audience. I ask myself that question constantly (and so should you!)  Amount of information and...

  4. Choosing the B(right)est Fluorescent Protein: Aggregation Tendency

    Type
    Blog Post
    ...between studies is substantial (e.g, the poor performance of mRuby2 is detected by Constantini et al (2015) ...selecting a bright fluorescent protein and a photostable fluorescent protein. In the third post of this...modules that fail to homodimerize in cells. To understand how well fluorescent proteins approach this ideal...dimerization tendency, as is present in EGFP. Constantini et al, (2012) developed a cell-based assay that...tagRFP does not act as a true monomer in cells (Constantini, 2012), whereas it was previously concluded to...chosen FP in your cell type of interest before starting your experiments.  Selecting a non-oligomerizing...CytERM with mTurquoise2 and dTomato, which are established monomeric and dimerizing FPs respectively, are...

  5. Plasmids 101: The Wide World of Natural Plasmids

    Type
    Blog Post
    ...organisms can’t use. Other plasmids offer resistance to toxic substances like heavy metals or pesticides, enabling... Blog Plasmids 101: Antibiotic Resistance Genes Antibiotic Resistance: An Old Solution but a New Problem...natural plasmids. Created with BioRender.com. Resistance (R) plasmids: microbial defense Genes allowing...its host. For example, cells with a penicillin resistance plasmid can grow in spite of a hostile Penicillium...fungus trying to kill them off. While antibiotic resistance-mediated selection is indispensable for cloning...cloning work, antibiotics and plasmid-encoded resistance to them are the tools of a millennia-old microbial...like colicin typically also carry the necessary resistance genes to help cells survive their own secretions...

  6. Plasmid-based Recombinant Monoclonal Antibodies: What They Are and Why You Should Be Excited About Them

    Type
    Blog Post
    ... for antibody expression. If starting from a protein sample (for instance, affinity purified antibodies...then be tested for antigen binding. If starting from an established hybridoma line, the antibody can be ...from a plasmid or from an integrated sequence in a stable cell line. The genes encode the heavy chain and...traditional hybridomas.  First, the long-term stability, consistency between batches, and molecular definition...and sequences for antibodies are not shared. For instance, through open plasmid sharing, the CRISPR community...antibodies, modify antibody function, improve antibody stability, and design tools that we haven’t even imagined...10.1038/521274a Begley CG, Ellis LM (2012) Raise standards for preclinical cancer research. Nature 483:531...

  7. Antibodies 101: Multiplex Immunofluorescence

    Type
    Blog Post
    ...combined in one step. As with standard IF, you will need to start with a fixed sample. Depending on... antibodies. The general process is similar to standard immunofluorescence, but with some extra considerations...photobleaching or by stripping out the antibodies, then start over with the next target: add the new primary, ... will need to include a marker like the Hoechst stain for the nuclei. (See Brewer et al, 2022 for a sample...Abcam's sample protocol is a good place to get started.  One important consideration when deciding whether...magenta), anti-AnkyrinG (green), and Hoechst nuclear stain (cyan). Panels C1-C3 show magnified detail of the..., many of the same considerations apply as for standard IF. However, there are a few specific considerations...

  8. Troubleshooting and Optimizing a Western Blot

    Type
    Blog Post
    ...forget to start it, the samples can actually migrate into the buffer — so it’s better to start over if ...consider using an unstained protein ladder, which will run a bit more true to size than a stained ladder. Unstained...transfer worked with Ponceau S staining or other reversible protein staining immediately after your transfer... Check transfer with Ponceau red stain or other reversible stain* Roll out all of the bubbles in...dilutions of your sample or protein of interest to understand the dynamic or working range of a given reagent... runs to optimize or troubleshoot.   Before you start… Mark your membrane! Membranes look the same from... amounts of protein, make sure your reversible staining method of choice is appropriately sensitive. If...

  9. Plasmids 101: Optimizing Plasmid Yields

    Type
    Blog Post
    ...time, plasmid prepping is a breeze. You get your stab from Addgene, streak for single colonies, sub-culture... genotype, as these mutations improve plasmid stability and yield. If a plasmid is prone to recombination...plasmids with inverted terminal repeats. We use NEB Stable to transform and prep high quality DNA that contains...recombination, but are also endA+. endA encodes a thermostable periplasmic endonuclease that can sometimes ...of common E. coli strains. Increase the volume Starting with a larger culture volume can improve plasmid...performing filter based plasmid prepping. Be sure to stay within the recommended range of the filter or you...growth conditions. For many high copy plasmids, standard LB broth and antibiotic concentrations will work...

  10. What's the Best Way to Elute and Store Your Plasmid DNA?

    Type
    Blog Post
    ...of preference, and it will be stable for years. Plasmid DNA can be stable at 4°C or even room temperature...answer: yes. This last step will influence the stability of your sample over time and determine which experiments...effectively use the DNA. pH and DNases affect DNA stability Figure 1: So many choices! What to do? ... The main factors that can affect DNA stability in solution are pH and DNase contamination.  Extreme... favors the breakage of the hydrogen bonds, destabilizing the molecule and favoring the separation of ... be able to store your sample without having substantial degradation and which experiments you can use... buffer is the best buffer for preserving the stability of your preparation for a long time. Tris buffer...

  11. 10 Ways to Share Your Science!

    Type
    Blog Post
    ...office is a good place to start. It’s not unusual to find science writers who started out as graduate students...skills. So, where can a graduate student or post-doc start? The good news is there are more opportunities than...Here are my recommendations on some ways to get started sharing your science with the community. Some of...father or your best friend who isn’t a scientist, start. If they don’t seem all that interested in the details... You’ll know you are on the right track if they start asking you questions. Remember that people like ...often, failure. It’s useful for non-scientists to understand not just the details of your science but also...written by practicing scientists. 3)   Consider starting up your own twitter feed or talking about and ...

  12. Antibody Validation for Flow Cytometry

    Type
    Blog Post
    ...have only been stained with secondary antibodies, showing the level of background staining. All antibodies... orthogonal approach to understand the relationship between antibody staining intensity by flow cytometry...different depending on the sample and target type. To start the validation process, one must first confirm that...permeabilized in 0.1% saponin. 400,000 cells were stained with the indicated Synaptotagmin-1 antibodies and...CoraLite® Plus 647 secondary antibodies. Antibody staining was quantified using the Attune NxT Flow Cytometer...Cytometer with representative images showing the staining intensity in the KO population (pink histogram)...maximize the separation and reduce any background staining. This can include using: different blocking reagents...

  13. Fluorescent Proteins 101: Green Fluorescent Protein (GFP)

    Type
    Blog Post
    .... Now GFP and its fluorescent derivatives are a staple in the lab. GFP is used in research across a vast...consisting of 238 amino acids derived from the crystal jellyfish Aequorea victoria. It has a fluorescent...that increased the fluorescence intensity and photostability of GFP. This also shifted its major excitation...to GFP and new iterations of fluorophores are constantly being engineered. Table 1 lists a few common ...Known Function S65T Increased fluorescence, photostability, and a shift of the major excitation peak to...Increases quantum yield for BFP Y145A and H148D Stabilizes the structure of Cerulean derivatives F99S,...GFP and other fluorescent proteins have become a mainstay in molecular biology. Scientists can easily utilize...

  14. Transferable Skills Guide: Teamwork

    Type
    Blog Post
    ...products of teams: Watson, Crick, & Franklin, Meselson-Stahl, Hershey-Chase...Who was I to dispute history? And... I happily embrace (and admittedly, I may have started that nickname myself). I have my own little Product...shipped; Chiara, the tech transfer expert who understands the somewhat complicated Material Transfer Agreement... kit pages. No single person could possibly understand every nuance of how kits work at Addgene, but...and others, we have updated our kit process substantially and have plenty of ideas for further improvements...biochemist, a geneticist, a microscopist, a biostatistician, and that one golden-handed technician who ...Andrew, the team’s founder, to create a mission statement in the first few meetings and we’ve volunteered...

  15. Tips for a 1st Time CRISPR User (by a 1st Time CRISPR User)

    Type
    Blog Post
    ... “You seriously didn’t add 3 microliters of 5 mM star anise?” This is something I was expecting when I...chicory root, but considering this is a needle in a haystack type of objective, it was surprisingly easy. It... See below for details. The entire process from start to finish took about 1 month. Cas9 delivery ...particular experiments, we generated A549 cells stably expressing Cas9 via transduction with Cas9 lentiviral...blotting was performed on whole-cell extracts from stable pools of A549 cells transduced with Cas9 expressing...with too many Cas9 integrations per cell, and my stable cell pool had wildly variable Cas9 expression (.../mL blasticidin for 9 days. Single cells from a stable cell pool were then expanded for an additional ...

  16. Antibodies 101: Avoiding the Mouse on Mouse Mess in IHC

    Type
    Blog Post
    ...are crisp, bright, and lacking in non-specific staining can be a challenge in the best of cases. But it... antibody, resulting in significant background staining (Figure 1A). Not all tissues will have the same...researchers can get away with species on species staining with no problem! But to be sure that your secondary...experiments. If you do find high levels of background staining by the secondary, then the following three strategies...help!     Figure 1: Species on species staining can be a problem because the secondary antibody...that you can significantly decrease background staining by mixing your primary and secondary antibodies...are a few common solutions to avoid background staining when doing species on species IHC. (A) One solution...

  17. Arabidopsis in Education: How the Arabidopsis Biological Resource Center Brings Plant Science to Life

    Type
    Blog Post
    ... Applied Plant Sciences at The Ohio State University. Established in 1991, the Arabidopsis Biological ...Arabidopsis and related species. Located at The Ohio State University in Columbus, Ohio, ABRC ships more than...Center (ABRC) set out to remedy that with the establishment of an outreach program in 2011. More than a...the Classroom, were developed and tested by ABRC staff. These kits provide in-depth, ready-to-teach lab...experiments allows them to develop a deeper understanding of scientific concepts than can be gained from...benefit of using Arabidopsis in the classroom is that staff from the ABRC are here to support you! From questions...guiding students through specific protocols, ABRC staff are available via e-mail and phone to answer questions...

  18. 15 Hot Plasmids from 2017

    Type
    Blog Post
    ...mCerulean3, mKOFP2) and empty vectors that have standard cloning sites or gateway compatible cloning sites...factors OCT3/4, SOX2, KLF4, and c-MYC is the established model for reprogramming somatic cells. A common...alkaline phosphatase-positive assay) as well as stabilization (Nanog-GFP reporter activation and silencing...like transgenic mice are important tools for understanding the function of cell types in the nervous system...they constructed AAV plasmids bearing one of the distal-less homeobox 5 and 6 (Dlx5/6) enhancer elements...stokes shift, long fluorescence lifetime, and a stable monomeric form as determined by the organized smooth...mScarlet-H with M164H (2-fold improvement in photostability compared to mScarlet, fluorescent lifetime ...

  19. Designing Your Chalk Talk for the Academic Job Interview

    Type
    Blog Post
    ...you as a colleague and give you a million dollar startup package. There are three steps to chalk talk preparation...what" question. For example, "we want a better understanding of the Z pathway" is vague and "what proteins...outcome makes it easier for the search committee to understand your goal, what you expect to find, and how you.... Finding evidence for or against a model is a standard scientific strategy. However, a robust research...Aims should have short titles, a clear easy-to-understand question/goal, an approach, anticipated outcomes...describing. It's easy to be overly ambitious – a common mistake is to make each aim into a 10-year research program...idea. It should be at a high level, broadly understandable to scientists outside your field, and interesting...

  20. 25 Work From Home Tips for Scientists

    Type
    Blog Post
    ... remotely. Social distancing? Many people are now calling this “physical distancing” since it’s important...important to stay virtually connected with the people in your life. At Addgene, we’ve started virtual activities...keep calm and science on. Practice self-care and stay connected Find time for exercise. Many studios ...these are activities that you can easily do while staying away from others.  Meditate. Meditating during ...lunch, and sharing photos of our pets. #SocialDistancing may send a wrong message for some people. .... We should recommend #PhysicalDistancing. Social interactions and #social support are even more necessary...your work online. Maybe you’ve always wanted to start using social media professionally but haven’t had...
Showing: 241 - 260 of 1052 results