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We narrowed to 957 results for: Cre

Showing: 361 - 380 of 957 results

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

    Type
    Blog Post
    ...Fluorescent Protein (GFP) has likely existed in creatures such as jellyfish for millions of years; however...physiological processes. Re-engineering GFP to increase its range of color and application Almost as.... Tsien described an S65T point mutation that increased the fluorescence intensity and photostability ...derivatives Mutation Known Function S65T Increased fluorescence, photostability, and a shift of the... the major excitation peak to 488 nm F64L Increased folding efficiency at 37C Y66W Causes the chromophore...Blue-shifts the wavelength (blue derivatives) Y145F Increases quantum yield for BFP Y145A and H148D Stabilizes...to identify particular cell populations in drug screens, to visualize micrometastases in nude mice in cancer...

  2. Antibodies 101: The Four ELISAs and When to Use Them

    Type
    Blog Post
    ...level of antigen in the sample. This figure was created with BioRender.com  Direct ELISAs A direct ... bind non-specifically to assay components and increase background staining. Direct ELISAs also lack the... antibody. Incorporating a secondary antibody increases the sensitivity of the assay, since multiple secondary... both primary and secondary incubation steps, increasing the workload and time commitment for users. This...antibodies for the capture and detection steps, which increases the specificity of the assay. The assay is also...antibody availability Longer protocolLess specificityIncreased chance of cross-reactivity Sandwich ... High specificityHigh sensitivity Longer protocolIncreased chance of cross-reactivityChallenging to develop...

  3. Plasmids 101: Broad Host Range Plasmids

    Type
    Blog Post
    ...they function in nature can inform us on how to create better recombinant vectors. In this blog post, ...provide tips on how we can harness those features to create broad host range recombinant vectors.  A plasmid... host range in plasmids? Not all plasmids were created equally. Only a handful of plasmids have been identified..., from broad host range plasmids can be used to create recombinant vectors that can replicate in different...vinelandii, Brevibacterium methylicum, Caulobacter crescentus, Desulfovibrio vulgaris, Erwinia carotovora, ..., Bordetella spp., Brucella spp., Caulobacter crescentus, Escherichia coli, Gluconacetobacter xylinus,...recombinant vectors can be modified further to increase the range of hosts in which proteins can be expressed...

  4. The PAM Requirement and Expanding CRISPR Beyond SpCas9

    Type
    Blog Post
    ...s lab performed a series of positive selection screens in bacteria to identify mutants of S. pyogenes ...sequences (Kleinstiver et al. 2015). From these screens, they identified four novel SpCas9 variants with...some cleavage with an NGA PAM. This variant may increase the specificity of genome modifications at DNA...allows for expanded PAM recognition as well as increased specificity and lower off-target activity (Hu ...proteins has and will continue to dramatically increase the number of CRISPR applications. The first non-Cas9...formation of “sticky ends” rather than the blunt ends created by Cas9. Cas12a  displays lower off-target editing...) system fuses Cas13b to RNA deaminase ADAR2 to create a specific RNA editor. Cas13 enzymes are advantageous...

  5. Designing Your Chalk Talk for the Academic Job Interview

    Type
    Blog Post
    ...significant knowledge gap in your field or even creating a new field. A knowledge gap is an impediment ...of experiments or a laundry list of molecules to screen has unclear utility. You cannot predict what will... have no hook. A battery of new assays for your screen result may or may not actually help. Ultimately...distinguish mechanistic questions from open ended screens and data collection. Organize your research vision...want to know the answer to your research problem. Creating a coherent unified story and model can help make...Consider going beyond the whiteboard. You may wish to create a sculpture or print a 3D model to better illustrate... His interests include the quality control of secretory proteins in the Endoplasmic Reticulum, optimization...

  6. Neuronal labeling with Spaghetti Monster

    Type
    Blog Post
    ...USA have used AAV1 and AAV2 serotypes in order to create an atlas of the mouse brain. This atlas enables...antibody recognition of epitope tags. Thus they have created antigenic protein tags called “spaghetti monster... FPs fused to multiple epitope tags. Indeed, to create the smFPs Looger’s team strategically inserted ...antibody-based amplification of their signals is weak and increases background fluorescence. Furthermore, many of ...instance, smFPs have been used to visualize “thorny excrescence” (TE) spines in the Hippocampal CA3 pyramidal...sites and primary antibody-based detection, smFPs create much brighter signals for weakly expressed proteins...: Making the journey from axon to nucleus AAVs CREATEd for gene delivery to the CNS and PNS Resources...

  7. To Codon Optimize or Not: That is the Question

    Type
    Blog Post
    ...Villarreal. By the late 1980s, scientists even created a codon adaptation index (CAI) that is based on... (Sharp and Li, 1987). In 2000 the Ikemura lab created the Codon Usage Tabulated from GenBank database...interest based on an organisms codon usage bias to increase translational efficiency and thus protein expression...across an entire polypeptide. Many researchers have created codon-optimization algorithms/resources (Athey ...study, codon optimization resulted in a 6-9 fold increase in expression of these channels providing scientists...to codon optimize? Although codon optimization increases protein production in certain systems, synonymous...optimization may not always be the optimal strategy for increasing protein production. If you are trying to express...

  8. Addgene Stands with the Scientific Community

    Type
    Blog Post
    ...limitations to facilities and indirect costs, and increased tariffs present an unprecedented challenge to ...to help research institutions maximize their increasingly limited resources: Preserving scientific assets...authenticated materials at a fraction of the cost of creating and verifying them independently — particularly..., and antibody production that labs may find increasingly difficult to maintain or access. These services...with industry and public and private partners to create additional pathways for critical scientific work...community has demonstrated remarkable resilience and creativity in the face of funding challenges. By strengthening...

  9. The 10 Most Distributed Plasmid Technologies in Addgene's First 10 Years

    Type
    Blog Post
    ...was Addgene's first distributed plasmid and was created in the lab of my thesis advisor, Dr. Bruce Spiegelman...requested item of 2005. shRNA - In 2006, we saw an increase in popularity of empty backbones for shRNA expression...requested. Ubiquitin - One particular gene we saw increased interest in from 2008 to 2009 was ubiquitin. The...popular with scientists, but 2012 was marked by increased deposits (and requests) of biosensors, optogenetics...and for so many different purposes that we had to create a CRISPR page to keep up with all the new plasmids...of plasmid sharing? Check out this infographic created by our own Addgenie, Laura Veckerelli. Click on...

  10. Antibodies 101: Normalization and Loading Controls for Western Blots

    Type
    Blog Post
    ...also helps confirm that lanes are loaded evenly. Created with BioRender.com. Loading controls are even...normalized values for the two conditions are equal. Created with BioRender.com. This kind of normalization...must be addressed with separate controls (green). Created with BioRender.com. Researchers often rely ... of the assay is between the two dotted lines. Created with BioRender.com. To see why this S-shaped...protein abundance versus band intensity (right). Created with BioRender.com. On this gel, the protein...loading control that can be used for normalization. Created with BioRender.com. Assumption #1: The loading...protein approach. By eye, it’s difficult to see an increasing concentration of β-actin in panel A. Indeed, ...

  11. Harnessing Bacterial Toxins for Allelic Exchange

    Type
    Blog Post
    ...Harvard. There’s an astounding number of ways to create chromosomal mutations in bacteria, so many that...of surrounding genes. However, there are ways to create scarless mutations, ones that don’t leave any undesired..., CRISPR-Cas9 technology has also aided in the creation of scarless mutations, both in stand-alone systems...systems (Jiang et al., 2013), and those which increase the efficiency of Lambda red (Pyne et al., 2015;... recombineering (Khetrapal et al., 2015). They created vectors for Lambda Red that contained one of eight...aeruginosa. These toxins were a mix of Type VI secreted effector toxins, toxin-antitoxin type toxins, ...Enterobacter cloacae, and Serratia marcescens and have created dozens of deletions as well as insertions with ...

  12. A new platform for fast functional characterization of potentially cancerous mutations

    Type
    Blog Post
    ...therapeutic interventions for cancer. This screening platform is an incredible boost to the cancer research community...Barcoding (HiTMMoB) technique (Tsang et al., 2016), creating a collection of plasmids each carrying a single...mutations The researchers used the collection to screen individual perturbations in vitro using Ba/F3 and...~22% of the mutations, however, significantly increased cell viability, and these “activators” may be ...annotation of rare gene aberration drivers of pancreatic cancer." Nature communications 7 (2016): 10500...

  13. What's New in CRISPR - March 2020

    Type
    Blog Post
    ... in yeast Base editing CRISPRi in Caulobacter crescentus Multiplexed gRNA for human pluripotent stem cells...editing. Using several rapid and cost-effective screening methods, David Liu’s lab identified and validated... base editor plasmids CRISPRi in Caulobacter crescentus Figure 1: Serial dilutions of CRISPRi in...adapted from Guzzo et al., 2020. Caulobacter crescentus is a popular model organism for studying the ...ctrA sgRNA and saw that the levels of ctrA mRNA decreased after 20 minutes in xylose (which controls expression...methylcytosine oxidation to DNA repair, which increases activation of methylation-silenced genes. Read...

  14. Technologies Enabled by NanoLuc® Luciferase

    Type
    Blog Post
    ...membrane depolarization increases the BRET signal whereas hyperpolarization decreases the signal. The authors...technologies that can accelerate your research. The creation of NanoLuc® Luciferase from a rather dull 19kDa... by NanoLuc® luciferase (NLuc) and set about to create an in vivo imaging reporter by linking two CyOFP1...-Lanterns with NLuc and called their improved creations Enhanced Nano-Lanterns. These great imaging tools...the two fluorescent proteins together thereby increasing their FRET signal. FRET is a non-radiative transfer...overlap with this optogenetic tool, they needed to create a calcium sensor with the following characteristics...

  15. Building Global Connections with the International Mentorship Program USA-EUROPE

    Type
    Blog Post
    ...International Mentorship Program (IMP) USA-Europe was created. IMP as an idea was conceived and developed by ...the specific technical and logistic support to increase the chances of success, but also nurture a supportive...country of origin with an international mentor creates a closer and more approachable relationship for...at IMP In these sessions, mentors helped mentees create “Career Development Plans” which provided paths...developmental needs and resources needed to pursue them, increased their network of contacts important for their ...Conference highlights included remarks by the Spanish Secretary of Research, Development, and Innovation, D. Carmen... as they witness their student’s progress and increases in self confidence. These improvements, combined...

  16. Fluorescent Proteins 101: Fluorescent Biosensors

    Type
    Blog Post
    ... biosensor creation, and custom FRET biosensors can be created using the cpFRET kit created by Olivier...Robert Campbell’s lab display calcium-dependent increases in fluorescence of up to 11,000-fold. FRET biosensors...biosensors. Blue flame “cameleon” calcium sensors created by Roger Tsien’s lab contain two fluorescent proteins...calcium binds, it triggers CaM/M13 interaction that increases FRET transfer between the fluorescent proteins...

  17. Early Career Researcher Toolbox: Free Tools for Making Scientific Graphics

    Type
    Blog Post
    ...are under a Creative Commons 3.0 license, which requires users to give appropriate credit, provide a link...about new scientific techniques and cool plasmids. Creating graphics were usually the last thing I thought..., I found some free design software that let me create graphics that better communicated the science in...reasons why this was the first program I used to create graphics for the Addgene blog. However, it has ...requested, although there’s no guarantee Biorender will create it. Additionally, you can upload your own images...

  18. ReFigure: Save Scientific Figures into Dashboards and Share Your Insights

    Type
    Blog Post
    ...experiments but we can definitely give you credit for it and increase your impact! Download ReFigure extension...along with the article metadata, legend and user created notes on a “Pinterest for scientists”? ReFigure...Linkedin and ResearchGate profile. To see how to create ReFigures in a few minutes, visit our YouTube [email protected] Please support ReFigure by creating ReFigures and answering this 2 minute survey! ...impact of teaching and doing science is rapid, concrete and open to innovation by all.  Additional Resources...

  19. Plasmids 101: Positive and Negative Selection for Plasmid Cloning

    Type
    Blog Post
    ...positive selection, negative selection, and/or screening. We’re focusing on positive and negative selection...in this blog post, but don’t worry, we’ll cover screens in a future post. Positive selection: Cells that...with or without your inserted DNA) and further screening will help you determine if the plasmid backbone...arises in yeast. For example, Markus Ralser’s lab created a yeast prototrophy kit containing 23 plasmids ...bacteria to grow on sucrose. Plasmid cloning: Creativity with naturally occurring genes  At this point... scientists are finding more and more ways to creatively perform selections in plasmid cloning. Additional...

  20. With Our New Viral Service, We're Taking CRISPR Further!

    Type
    Blog Post
    ...CRISPR pooled gRNA libraries that can be used to screen for genes involved in a wide variety of cellular...processes. While these are widely requested and incredibly useful tools, they require extensive background... infect target cells, and, finally, conduct the screen. To help alleviate some of this labor for our ...optimization-scale infection and at least one screening-scale experiment achieving the gRNA representation...) says: "When you're setting up to do a genetic screen, you need to focus on building a really good model...use along with lentiviral packaging plasmids to create your own lentivirus stocks. Our research scientists...
Showing: 361 - 380 of 957 results