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We narrowed to 971 results for: lars

Showing: 361 - 380 of 971 results

  1. Antibodies 101: Affinity Reagents

    Type
    Blog Post
    ...reagents.  Affinity Reagents  Affinity reagents are molecular tools that have the ability to specifically recognize...affinity reagents is their size. Antibodies are large enough that their bulk can be a barrier at times...difficult to source, may not be readily available for a large number of targets, and can require specific conjugates...   Types of Affinity Reagents  (Listed in no particular order.) Fab fragments are created through enzymatic...variables (scFvs) are ~27kD engineered fusion proteins similar to a Fab fragment but created through engineering...eukaryotic cells, allowing for them to be used as intracellular inhibitors.  Aptamers are not proteins at all...

  2. Antibodies 101: Fab Fragments

    Type
    Blog Post
    ...described above can be used for similar applications and have similar advantages, though with some caveats...chain. Created with biorender.com     The modularity of antibodies means that it is relatively easy..., two full antibodies bound together may be too large. Using a complex that is one full antibody bound...Alternatively, Fab fragments can be used as a block in similar scenarios. By applying unconjugated Fab fragments...and have lower binding affinity than some of the larger fragments.  Think of antibody fragments as new ...

  3. Plasmids 101: CcdB - The Toxic Key to Efficient Cloning

    Type
    Blog Post
    ... we will focus on a unique gene that makes the popular GatewayTM method possible: ccdB. But what is ccdB...start with a brief history of the gene and how molecular biologists have harnessed it to evolve cloning...clones.   ...Becomes a powerful cloning tool Molecular biologists first saw the potential of this system...and are considered F-); however, there are a few popular lab strains, such as NEB Stable, JM109, XL1 Blue...experiments. Please check out Addgene’s Choosing a Molecular Cloning Technique page and the Addgene Blog for...Cloning Posts Resources on Addgene.org Choose a Molecular Cloning Technique Find Plasmid Cloning Protocols...

  4. A New Generation of Adenine Base Editors Improves Editing in Primary Human Cells

    Type
    Blog Post
    ...and TadA7.10, they designed one set of ABE8’s similarly (ABE8.x-d). In order to assess the effect of removing...you need to modify multiple genes to generate a cellular phenotype. Again ABE8.20-m was the winner. It ...editing in ABE8s While base editors are precise molecular tools, they do have undesired off-target editing...have been described to introduce base changes in cellular DNA and RNA dependent or independent of the gRNA..., and a thoughtful selection of guideRNAs. Of particular note, ABE8s reported in this work did not cause...sites that used to be difficult to target. In particular, their ability for more robust editing in human...

  5. Synthetic promoter AAVs for cell-type specific expression in retinal cells

    Type
    Blog Post
    ...genes in particular tissues, but they are costly and time-consuming to generate, especially for large animals...Adeno-associated virus (AAV) is a popular tool for gene delivery, but it has a drawback: how do you ensure... it to? Knowing that a gene is expressed in a particular cell type is important not only for translational...types.  Botond Roska’s lab at the Institute of Molecular and Clinical Ophthalmology Basel recently took...Besides restricting AAV transgene expression to particular cell types or classes, these synthetic promoters...

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

    Type
    Blog Post
    ... to interact by stimulation with blue light.  Similarly, light can also be used to control homo-oligomerization...distribution of clusters without accumulation in subcellular compartments such as nuclear speckles. Furthermore...OptoSTIM1), caused two-fold faster changes in intracellular Ca2+ levels than the original version containing...CRY2clust should prove a useful tool to further probe cellular function. The CRY2clust Plasmid can be found here... commonly observed with direct stimulation are largely absent in the soCoChR-expressing system. Taken ...interactions. We’re excited to see how you adapt similar FLINC biosensors to work in your lab. Find the ...

  7. Corralling and Cornering the Goldilocks Antibody

    Type
    Blog Post
    ... very specific biological targets. However, as popular and useful antibodies are, they are in a midst ...as non-human primates and humans. Since we need larger-volume antibodies for thicker tissues and whole... immunoglobin component that can be produced in large “copies.” This homogeneity circumvents the batch-to-batch...reproducibility and cost-effectiveness for broad and large quantity uses. We therefore use both rAb and mAB...same target to see if the results are the same/similar, emphasizing the antibody’s specificity. Strength...from the background noise. If the sample images similarly to the negative control, that informs us that ...

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

    Type
    Blog Post
    ...encode proteins. They play important roles in many cellular and regulatory processes, but much of their functionality...removing the full ncRNA locus (Ho et al. 2012). For large-scale screening, sgRNA libraries are useful for ... same time (Liu et al. 2018). However, removing large lncRNA fragments with multiple gRNAs poses the risk...Zibitt et al. 2021). Mutating splice sites is particularly effective as it can result in the retention ...Mutations of splice sites. (C) Removal of an exon or a large genomic fragment. (D) Knock-in: insertion of a synthetic...used to target lncRNAs with high specificity, particularly in applications requiring smaller delivery systems...

  9. Making CRISPR Plasmids Using Fragmid

    Type
    Blog Post
    ...developed third option: Fragmid. Fragmid is a modular vector assembly process designed for a quick, plug-and-play...vectors in five days. Did you say… easier? Fragmid’s modular assembly structure takes a lot of the design work... And it truly is plug-and-play. Because of its modular cloning approach, it’s easy to switch out reporters... by phiC31   The Fragmid website While the modular cloning approach is key to the Fragmid system, ..., M. A., Green, T. M., & Doench, J. G. (2024). Modular vector assembly enables rapid assessment of emerging....2024.100519 More resources from Addgene Plasmids 101: Modular Cloning Addgene's CRISPR Guide Plasmids 101: Golden...

  10. Plasmids 101: Terminators and PolyA signals

    Type
    Blog Post
    ...polyadenylation signal by using in vitro analyses. Molecular and Cellular Biology. 1992. PMID: 1333042. PubMed Central... thought that poly(A)s are used to control the cellular concentration of regulatory RNAs and may additionally...secondary structure to induce transcript cleavage, similar to the Rho-independent termination found in prokaryotes...terminate transcription. Polymerase I uses a process similar to the prokaryotic Rho-dependent mechanism, whereas...Terminators: The RNA 3′-End Chronicles. Journal of molecular biology. 2011. PMID: 21439297. PubMed Central ...

  11. The Materials Science of Optogenetics Experiments

    Type
    Blog Post
    ...Simon! Derek P. Simon received his PhD in Cellular and Molecular Biology from the University of Michigan...blogger Derek Simon. The surgeries and standard molecular neuroscience validation experiments we discussed...cemented to the animal’s skull using dental cement (a similar procedure as implanting a guide cannula). A fiber... Gi/o protein-coupled vertebrate rhodopsin in cerebellar Purkinje cells. The Journal of biological chemistry...PMCID: PMC3466483. 4. Kim TI, et al. Injectable, cellular-scale optoelectronics with applications for wireless...

  12. No Llamas Required - Synthetic Nanobodies Against Membrane Proteins

    Type
    Blog Post
    ... that can bind effectively to protein targets, similar to how antibodies do. The structure of the common...(~150 kDa) and Y-shaped because it contains two large heavy chains and two small light chains that stick...antibodies to get to, like ones that are hidden in the molecular crevices proteins on the cell surface. Figure...optimally binding sybody, you want to start out with a large and diverse pool of sybodies that hypothetically...because it has the advantage of starting out with a larger initial starting library size (up to 1012 members...Characterization of Enzyme Variants. In: Methods in Molecular Biology. Humana Press, pp 133–148. https://doi.org...

  13. Adenoviral Delivery of CRISPR/Cas9 Aims to Expand Genome Editing to Primary Cells

    Type
    Blog Post
    ...already a packaging capacity larger than AAV vectors. This capacity is large enough to carry the Cas9 gene...compared to other viral vectors, AdVs can provide a large transgene packaging capacity. For instance, with...non-transformed cells yielded rates of targeted mutagenesis similar to those achieved by isogenic AdVs encoding TALENs... B-thalassemia, Sickle Cells Disease, Duchenne Muscular Dystrophy, Alpha1-antitrypsin deficiency (most... animals in order to create animal models for particular cancer. That’s exactly what the lab of Andrea...

  14. Quick Guide to Working with Drosophila Part 1: Getting Started with Flies

    Type
    Blog Post
    ...into embryos and then become larvae. There are three larval stages where larvae molt in order to continue...continue growing in the next larval stage. After the third larval stage, Drosophila will pupate, which takes...fertilized egg by its mother. Inside the pupal case, the larva will metamorphose into its adult form. In all, it...

  15. Production of Virus in Insect Versus Mammalian Cells

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    Blog Post
    ...cells are a great option for viral production, particularly for labs that already do mammalian cell culture...for the generation of viruses, though they are a popular choice for protein production. The most commonly...infect insects. This is what makes insect cells a popular choice for protein production. AAV and lentiviruses...systems. Baculovirus transfer plasmids have a much larger packaging capacity — up to 38 kb! Generally, insect...in between passages to eliminate dead cells and cellular debris. While adding time (about three to four...supernatant in a centrifuge to remove any unwanted cellular debris, as you would when collecting a lentivirus...

  16. Plasmids 101: Knockout/Knock-In Plasmids

    Type
    Blog Post
    ...specific genes in mammalian cells. Despite the popularity of CRISPR-based knockout/knock-in systems, these...nucleotide sequences are exchanged between two similar or identical molecules of DNA. Gene targeting takes...targeting constructs. In the example shown in Figure 1, large sequences corresponding to exons 1 and 3 of the ... the cells that have randomly integrated all or large portions of the plasmid. Deliver your construct ...insert GFP into the genome, we’d create a plasmid similar to the one shown below, where the sequence of GFP...CRISPR is very good at making knockouts, knocking in large sections of DNA can be more difficult. Addgene depositors...

  17. Plasmids 101: Optimizing Plasmid Yields

    Type
    Blog Post
    ... might need to use multiple techniques for a particularly stubborn prep. Check the bacterial strain Increase...strains, including HB101 and derivatives, release large amounts of carbohydrates that can inhibit the cell...levels of protein, not plasmid DNA. If you need large amounts of DNA from a plasmid in a protein expression...choice of strain will ultimately depend on the particular features of your plasmid, it is advisable to ...coli strains. Increase the volume Starting with a larger culture volume can improve plasmid yield. Our lab...gas exchange in the overnight culture. Generally, large cultures in flasks may be shaken around 220 RPM,...

  18. Anatomy of a Plasmid Page at Addgene

    Type
    Blog Post
    ...depositing lab. There may be other plasmids with similar names in the collection, but you can use this name...some of Addgene’s plasmids is an indication of popularity. A yellow flame indicates that a plasmid has ... on our website, one of the available filters popularity (e.g. only showing those plasmids which have ... than 100 times).     I guess it's cool to be popular, but how do I know if a plasmid is actually useful...If the depositor has indicated that there is a particular mutation found within the gene/insert, this information...means you can see which labs have already used a particular plasmid, and which experimental systems it was...

  19. Genome-wide Screening Using CRISPR

    Type
    Blog Post
    ...role of individual genes or entire pathways in a particular disease process. While CRISPR is certainly not... mutant cells to identify genes involved in a particular phenotype. CRISPR can be readily scaled up for...fluorescence-based cell sorting and screened for a particular phenotype. For example, mutant cells can be used...cell types that are of low abundance are not particularly well suited for genome-wide screening. Avoid...targeting the same gene can be strong evidence that a particular gene is actually important for a given phenotype...Lentiviral CRISPR Libraries. Current Protocols in Molecular Biology 115: . https://doi.org/10.1002/cpmb.8 ...

  20. Tips for Using BLAST to Verify Plasmids

    Type
    Blog Post
    ...results. The default setting is “Optimize for Highly similar sequences (megablast)”, which is very fast and ...often unreliable, we suggest using the “Somewhat similar sequences (blastn)” program for blastn so that ...your sequencing trace file. Unlike megablast, the regular blastn program uses a smaller word size and lower...protein sequences (nr)” database as it contains the largest number of protein sequences. Just below the BLAST...options and for our suggested blastx customization. Similar to nucleotide sequences, proteins often have repeated...COBALT for aligning multiple protein sequences, particularly for comparing different isoforms. In addition...
Showing: 361 - 380 of 971 results