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Showing: 181 - 200 of 755 results

  1. Tagging Optogenetics and Chemogenetics Receptors: Fluorescent Proteins and Other Options

    Type
    Blog Post
    ...method to identify the expression of transgenes of interest. Furthermore, fluorophore variants (for example...simultaneous identification of more than one protein of interest. However, in our experience, we found that fluorescent...than an aid, in the expression of the protein of interest. The green fluorescent protein (GFP) and mCherry...interfere with the correct function of the protein of interest. In our studies, we used AAV viral vectors to ...influence on the expression of the transgene of interest and its localization in the membrane. The lesson... can impact the expression of the transgene of interest. In our studies, mCherry interfered with the transport...could help improve the transport of protein of interest the membrane.   Figure 1: Electron microscope...

  2. The Scientific Conference Poster Session: Tips for Success

    Type
    Blog Post
    ...including more background on how your virus of interest causes cancer, as well as therapeutic strategies...the conference, map out which posters you are interested in viewing. For most larger conferences, poster...session, so bookmark which posters are of most interest to you a few days in advance. If you don’t have...If you come prepared with a list of posters of interest, then you’ve already set yourself up for success... – kindly let the presenter know you are very interested in his/her work, and would enjoy hearing the ...constraints, there may be particular poster you are interested in viewing which has drawn a large crowd, making...Time permitting, you can go back to the most interesting and popular posters at the end of the session...

  3. Scientist Networking: What is an Informational Interview?

    Type
    Blog Post
    ... do during your training is meet interesting people doing interesting things. I call this building relationships...learning new things from other scientists doing interesting work. Consider this to be like any other research...research project. You’ve met someone whose career interests you or you want to pursue someone doing a job ...nonthreatening request to discuss experience, interests and career path. Because there is no demand for...what networking is about), but be upfront about interest in a job. Ask openly to be connected to someone...people like to help One good place to look for interesting connections is your university’s alumni database...too busy to respond to you). You will uncover interesting opportunities by meeting with people working ...

  4. Viral Vectors 101: Systemic Capsids

    Type
    Blog Post
    ...regions. If you’re interested in studying the brain in a way that goes beyond such restrictions, you may want...Mich et al., 2021) can help restrict expression to your cell type of interest, while miRNA target sites ...large or diffuse biological structures. Of primary interest here are systemic capsids that target the central..., including species, strain, age, sex, area of interest, and health. While it’s always helpful to start...your average AAVs — and have many of the same restrictions, like limited cargo space (~4.7 kb) for your...used to prevent expression in cell types not of interest. Systemic capsids packaging Cre-dependent transgenes... with Cre-expressing transgenic mouse lines to restrict transgene expression to specific tissues and cell...

  5. Four Base Editing Reporters to Monitor and Enrich Editing in Real-time

    Type
    Blog Post
    ...Harris Lab relies on the restoration of a frame-shift in mCherry that restores fluorescence to monitor ...that disrupts fluorescence due to a frameshift. Restoration of fluorescence can only occur through editing...results in two DSBs that can be repaired by NHEJ restoring mCherry expression. Cells can then be sorted using...C-to-T editing by the APOBEC-Cas9 editosome would restore eGFP fluorescence, making it easy to quantify single... point mutations in eGFP that results in the restoration of fluorescence. Transient reporter for editing...-GFP conversion and to target specific loci of interest in the genome. The Wang and Brafman lab showed...higher frequency of genomic base pair edits of interest compared to cells segregated solely by a reporter...

  6. FastCloning: A Newer, Simpler Cloning Technique

    Type
    Blog Post
    ...Comparison Between Standard Restriction Cloning and Fast Cloning Requirement Restriction Cloning Fast Cloning...elements of biomedical research. Traditional restriction cloning is one of the oldest of these techniques...purification, ligation, and transformation. While restriction cloning is still routinely performed by many ...Cloning Choose Restriction Enzymes + - PCR Amplification + + Digestion + + Gel Purification + - ...products and digest with methylation sensitive restriction enzyme DpnI to remove the templates. Digestion...graduate student at the University of Connecticut interested in protein science.     References: 1. Celie,... restriction cloning...

  7. Antibodies 101: Introduction to Gating in Flow Cytometry

    Type
    Blog Post
    ...cells of interest do not lie beyond the dimensions displayed in the plot. (B) A more restrictive gate results...of my cells of interest lies within a clear negative population. When you’re interested in measuring a... to select and precisely measure your cells of interest. In many experiments you’ll be working with a ... on the following gates, cells that are not of interest can still be gated out later. Based on personal...(single cells, live cells, my marker stains of interest). Let’s double-click inside our gate and proceed...you have identified and gated on your cells of interest, you will likely need to exclude all duplicate...multiple individual gates around the populations of interest (Figure 6). A.                                                                                               ...

  8. Using Phosphoserine to Study Protein Phosphorylation

    Type
    Blog Post
    ...cloned with an ORF of interest for customized experiments. Once the ORF of interest is cloned into one of...phosphomimetic residues. The phosphorylated residue of interest is mutated to a negatively charged residue, aspartate...plasmid encoding the open reading frame (ORF) of interest (Figure 2). 1. The phosphoserine OTS The backbone...a plasmid encoding their open reading frame of interest. Two expression systems have been verified for...instead of a glutamate which allows the system of interest to be validated first through the production of... generate as much pSer-incorporated protein of interest as possible, some optimization will be necessary...to a single phosphate moiety on the peptide of interest.  After confirming pSer incorporation, researchers...

  9. Extracurricular Activities for a Strong Science Career Path

    Type
    Blog Post
    ... and interests. Are you particularly good at writing and editing manuscripts? Are you interested in scientific...science professions based on their skills and interests. Once scientists identify potential career pathways... academic setting and to shape and strengthen interests and skills. Although the word “extracurricular... potential careers that suit these skills and interests: Use career planning tools such as the Individual...opportunities beyond academia. Once a profession sounds interesting, TAKE ACTION! Taking action: Examples of extracurricular...outside your particular field of study Assess your interest in a different field Outreach activities...information about shaping your career to your values, interests, and skills? Curious to know where graduate students...

  10. 27 Hot Plasmids from 2016

    Type
    Blog Post
    ...unique restriction enzyme site and unique primer sites for more rapid screening by restriction digest ..., et al. Cell Rep. 2015. PubMed PMID: 26190106 PRESTO-TANGO: An open-source resource for drug discovery...ligands for orphan GPCRs is of significant medical interest, however efforts to do so have been hindered by...receptors. Bryan Roth's lab has a newly available PRESTO-TANGO kit which allows researchers to interrogate...publication, Kroeze et al. describe their enhanced TANGO arrestin recruitment assay incorporating 314 codon-optimized...and Screening via Transcriptional Output-TANGO (PRESTO-TANGO) to screen the NCC-1 library of approved ...discovery. Researchers can use plasmids from the PRESTO-TANGO kit directly for drug screening or easily...

  11. PiggyBac-ing Through the Genome Editing Field

    Type
    Blog Post
    ...efficiencies that may be low in the cell line of interest. The piggyBac™ system allows for significantly...of the donor-transposon (carrying your cargo of interest) into random ‘TTAA’ sites throughout the genome...line-dependent, and should be tested in the cell line of interest. Importantly, by varying these ratios, you can... or minimize expression levels of your gene of interest, as needed. Controlling the expression levels ...isolating the specific cells that have the edit of interest. Fortunately, by combing nuclease technology with...transposon in a donor plasmid carrying the DNA edit of interest, along with a site-specific nuclease (such as ...thousand clones to find a cell line with the edit of interest. With the larger piggyBac™ donor vector, it is...

  12. Antibodies 101: Secondary Antibodies

    Type
    Blog Post
    ...protein, and then voila! Wherever your protein of interest is, a signal will also be. This is the direct ...giving you enough signal to detect your protein of interest, it may be time to look at the indirect approach...While a primary antibody binds to the protein of interest, a secondary antibody binds to the primary antibody...molecules associated, indirectly, with each protein of interest.    Figure 1: The direct approach (left...secondary antibodies are indicated when the protein of interest is found in low amounts or if the conjugated signaling...primary antibody for your assay and protein of interest is often one of the most challenging components...multiplex assay - one with multiple proteins of interest - the secondary antibody needs careful consideration...

  13. A Practical Guide to Optimizing AAV DIO and FLEx Vector Expression

    Type
    Blog Post
    ...recombinases like Cre or Flp in their cells of interest and recombinase-dependent AAV vectors for selective...to achieve adequate expression in your cells of interest.  The AAV serotype used can exhibit profound differences...you are using transduces cells that you are not interested in with greater efficiency, it could lead to ...understand which serotype best transduces your cells of interest. We recommend testing several different serotypes...determine which one is the best for your cells of interest.  Determine injection titers and volumes based...injection parameters needed to transduce your cells of interest. Using a constitutively expressing vector for ...is important to test it in the brain region of interest, especially when using DIO/FLEx vectors for off-target...

  14. Plasmids for Endogenous Gene Tagging in Human Cells

    Type
    Blog Post
    ...precise location of the protein or structure of interest. Endogenous gene tagging is an ideal solution ...creating the plasmids was to identify the gene of interest and decide which terminus, N or C, to tag and ...researchers who study the protein or structure of interest. For a C-terminal tag, as shown in the example...contains 1 kb homology arms matching the gene of interest, a linker, and the GFP tag sequence to be inserted...near either the beginning or end of the gene of interest —in our case in the genome of an hiPS cell. While...this example, at the C-terminus of the gene of interest. To get the molecular components of CRISPR/Cas9...plasmids for introducing tags into your gene of interest. Researchers have developed a variety of endogenous...

  15. Antibodies 101: ELISA (Enzyme-linked Immunosorbent Assay)

    Type
    Blog Post
    ...antibody that is specific to the protein you are interested in detecting in your sample. When an ELISA kit...wells, and the antibodies bind to your protein of interest.  The next step is to detect the protein-antibody... sample. However, in this case your protein of interest is pre-incubated with a known amount of an antibody...antibody (meaning it hasn’t bound to your protein of interest) will bind to the competing antigen in the wells...inverse measure of the amount of your protein of interest in the original sample. The competitive ELISA ...sandwich or competitive ELISA because the protein of interest is then detected by an enzyme-conjugated antibody...used to calculate the amount of your protein of interest that is present. Using a Standard Curve To do ...

  16. Technical Design of a Western Blot

    Type
    Blog Post
    ...impact the antibody’s binding to your protein of interest. Protein preparation Before you can blot your ...incubation temperature for your specific protein(s) of interest.  Determining linear detection range If you are...depends mostly on the weight of the proteins you’re interested in. Table 1 lists the most common types of gels...in a variety of pore sizes. If your protein of interest is small, you may want to consider a smaller pore... size (0.2 µm). Conversely, if your protein of interest is large, look for a larger pore size (0.45 µm... and are worth considering if your proteins of interest vary wildly in size. However, these systems are...It’s fairly easy to decide!  If your protein of interest is available in pure form, you can use it to create...

  17. Fluorescent Proteins 101: GFP Fusion Proteins - Making the Right Connection

    Type
    Blog Post
    ...University of Amsterdam). Tagging a protein of interest with a fluorescent protein to study its function...all of the native properties of the protein of interest can be challenging. In this blog I discuss strategies...relatively long linkers between the protein of interest and the fluorescent protein. These were either...fusions at the N- and C- termini of your protein of interest The most straightforward fusion protein is made...C-terminus of the fluorescent protein and protein of interest (POI). To indicate the sequence, people often ...mScarlet-tubulin (Plasmid #85045). We often use restriction enzymes in the multiple cloning site for generating...Inserting a fluorescent protein within your protein of interest We have encountered several proteins for which...

  18. Antibodies 101: The Basics of Western Blotting

    Type
    Blog Post
    ...ve gotten the plasmid encoding your protein of interest from Addgene, transfected it into your target ...lane can help confirm the size of the protein of interest. An electric current applied to the gel causes...with a primary antibody against your protein of interest. But which antibody should you use? You will likely...different antibody options targeting your protein of interest from a variety of vendors and you may even find...expression knocked down or knocked out. If you are interested in making your own knockout line for validation... Bauer's labs.    Visualizing your protein of interest with the help of secondary antibodies In order...the primary antibody, which binds the protein of interest, is not labeled. Instead, a conjugated species-specific...

  19. How to Make Friends and Meet People at a Scientific Conference

    Type
    Blog Post
    ...more easily. Attend special interest group sessions that match your interests. I favor small conferences...provide the opportunity to meet people who are interested in the same things you are on a deep level. Right...common. Namely, the scientific question you are interested in and this is a great ice breaker. Of course...relationships go further and grow over time, but being interested in the same phosphate of your favorite kinase...natural time to meet someone since you are both interested in the same talk. Read more on this advice to...say so. If you read a paper that you think would interest them, send it and ask for their input. Connect...

  20. Viral Vectors 101: An Introduction to AAV

    Type
    Blog Post
    ...gene of interest (or genes) need to be 4.7 kb or less to fit in a single rAAV. For those interested in heftier...icosahedral capsid, that can be used to express genes of interest in cell and animal models. AAVs are non-pathogenic...host cell. In order to make room for a gene of interest, AAVs used in research have their genomes “gutted... than in vitro. On the other hand, if you’re interested in expressing multiple small genes, you can design...expression via switches placed in your cell type of interest. Careful combination of promoters, gene switches...legend from Okada, 2013.   scAAV If your gene of interest is truly small (<2.5Kb) and you need expression...
Showing: 181 - 200 of 755 results