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We narrowed to 1,050 results for: ins;

Showing: 281 - 300 of 1050 results

  1. Open Resources and Plasmid Tools For Studying C. elegans

    Type
    Blog Post
    ...sharing their worm strains through the Caenorhabditis Genetics Center (CGC) which maintains and distributes...each half to a gp41-1-N-intein. Split inteins are protein domains that naturally “associate, self-excise...distributes the strains all over the world. When I was at the 22nd International C. elegans meeting, I was again... the Arizona State University (ASU) Biodesign institute developed a 3’UTR database specifically for C.... discovered in plants to conditionally deplete proteins of interest in C. elegans (Zhang et al., 2015)...modified TIR-1 protein can degrade degron-tagged proteins in an auxin dependent manner. This system allows...scientists to both spatially and temporally knock-down proteins of interest.  Plasmids for cGAL and split cGAL...

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

    Type
    Blog Post
    ...long ncRNAs is impractical. Instead, tags or stop elements can be inserted to disrupt the transcription...RNAs (ncRNAs) are transcribed but do not encode proteins. They play important roles in many cellular and...permanently disable, protein-coding genes by generating insertions or deletions in the DNA sequence. This same approach...break works well because very small deletions and insertions in the DNA sequence can completely disrupt expression... an open reading frame, so small deletions or insertions may not result in a functional loss (Ho et al...the ncRNA (Hazan and Bester 2021). For example, inserting a polyadenylation signal downstream of the transcription...exon or a large genomic fragment. (D) Knock-in: insertion of a synthetic polyadenylation (spA) signal. Figure...

  3. Hot Plasmids February 2024

    Type
    Blog Post
    ...AAV.CAP-Mac for systemic gene transfer in primate brains Endothelial-specific AAV delivery Recombinant anti-FLAG...COX8a-mStayGold. Scale bars 10 μm (main), 1 μm (inset). Image reused from Ando et al. 2023, under CC-BY...for your next experiments, they will also likely inspire further-improved versions. Thanks to Christophe...Leterrier, whose FocalPlanes piece on mStayGold inspired and informed this section.  Find mStayGold plasmids...2023). Bright and stable monomeric fluorescent proteins derived from StayGold. ResearchSquare (Preprint...alchemy: CAP-Mac unleashes the secrets of primate brains with brilliance By Hridaya Patel (Addgene co-op...systemic gene delivery in non-human primate (NHP) brains.  Systemic gene delivery capsids previously identified...

  4. Antibodies 101: Antibody Engineering and Directed Evolution

    Type
    Blog Post
    ...families of cell surface and secreted proteins (glypicans, integrins, and more coming soon) as well as epitope...engineer the most miniscule parts of life, like proteins, to study or alter their function for a variety...Degenerate primers are a group of primers that contains a variety of degenerate codons (different codon...amino acid) for various amino acids. Essentially, instead of forcing sequence diversity using error-prone...guided directed evolution might be more your style. Instead of leaving sequence generation to chance, you start...regions you want to evolve. With this added layer of insight, you can shrink your mutant library and increase...binding specificity, you can perform binding assays against the target antigen to identify mutants with enhanced...

  5. Our Most Popular Blog Posts from 2019

    Type
    Blog Post
    ...PS: they’re free. 3. Fluorescent Proteins 101: GFP Fusion Proteins - Making the Right Connection by Joachim...optogenetics, chemogenetics, plasmids, fluorescent proteins, careers, and more. These resources can now be...Fluorescent Voltage Indicators, and Photoswitchable Proteins by Cary Valley, Shreya Vedantam, Michelle Cronin...used in cloning. 8. Bright Monomeric Fluorescent Proteins: mNeonGreen, mTFP1, and mWasabi by Jennifer Tsang...and find plasmids containing these fluorescent proteins at Addgene. 7. Designing Your Chalk Talk for ...Fluorescent Vegetables in Aptamer Soup by Aliyah Weinstein RNA aptamers, which are nucleic acid sequences...shares some strategies for generating GFP fusion proteins. From considering protein size and shape, linker...

  6. Tips from the Repository Trenches: Using Barcodes to Track Samples

    Type
    Blog Post
    ...plasmid's journey begins First, each package is opened and the contents are carefully inspected to verify that...different strains of E. coli that can be used for transformation. The preferred E. coli strains for storage... then the journey of transformation and storage begins. Some samples are submitted as bacterial colonies...the year (see graph below). Addgene scientists inspect each sample and make sure they have been barcoded...assigned to a location in a metal freezer rack that contains five levels that can hold five plates on each ... back into because every barcode on every tube contains information about that tube’s appropriate location...been received and stored, it is put into E. coli strains that have been treated to easily take in plasmids...

  7. Antibodies 101: Designing Your First Flow Panel

    Type
    Blog Post
    ...the surface or the inside of your cells. While proteins themselves can emit intrinsic fluorescence when...amino acids found in all proteins, so you can't distinguish the different proteins from each other. To distinguish...Lastly, there are fixable dye stains that react with free amines on and inside the cells and are thus typically...distinguish different target proteins in your flow analysis, you'll need to use fluorophores that attach...appropriate readout. A common method of labeling proteins is to fuse them with a fluorescent reporter, such... reporter's fluorescence. Another way to label proteins is through treating cells with fluorophore-conjugated...antibody-fluorophores, fluorescent reporters, and/or dye stains used in your flow cytometry experiment are known...

  8. A Primer on Optogenetics: Introduction and Opsin Delivery

    Type
    Blog Post
    ... of Antonello Bonci, Director of the National Institutes of Drug Abuse Intramural Research Program (NIH-NIDA...are numerous variants for the different types of opsins available and their differences and uses have been...discussed thoroughly elsewhere (here and here for instance) [2], [3]. Karl Deisseroth of Stanford University...details about types of constructs available, instructions for obtaining a material transfer agreement,...published at this point.  In brief, some standard opsins (see table above) include Channelrhodopsin2 (Chr2...and subsequent firing of an action potential. Halorhodopsins (HR) are green/yellow-light sensitive chloride...specific opsin expression. However, the types of opsins you are able to use are obviously limited by the...

  9. CRISPR Activation: A Practical Guide

    Type
    Blog Post
    ... Chavez, researchers at the Wyss Institute for Biologically Inspired Engineering. CRISPR/Cas9 is an enormously...and former researcher at the Wyss Institute for Biologically Inspired Engineering.     Alejandro (Alex)...prevent your gene from being up-regulated. For instance, generally, the more highly expressed a gene is...similar tools were made using other DNA binding proteins such as zinc fingers (ZF) and TAL effectors (TALE...scale CRISPR-based screens can be used to gain insight into protein structure and function (Pan et al....Ansari R, Harper S, Jackson DA, McRae R, Pooley R, Wilkinson P, van der Meer D, Dow D, Buser-Doepner C, Bertotti...Davidsohn N, Housden BE, Perrimon N, Weiss R, Aach J, Collins JJ, Church GM (2015) Highly efficient Cas9-mediated...

  10. Identifying Sequence Elements with SnapGene's Feature Database

    Type
    Blog Post
    ...microbial strains. It proved to be impractical to catalog every variant of a feature. Instead, we identified... days or even weeks. Primer design was done painstakingly by hand. Records of plasmid construction were...their goals as intuitive and painless as possible. Instead of crowding the interface with every possible option... antibiotic resistance markers and replication origins, but there is extensive heterogeneity in the feature...efforts, such an algorithm has limitations. For instance, it could miss a common feature if the sequence... inaccurately. The best example is fluorescent proteins, which often come in closely related versions ... genomes of newly sequenced bacterial or viral strains. We are working on a way for SnapGene users to ...

  11. Antibodies 101: Chimeric Antibodies

    Type
    Blog Post
    ...and find an ideal antibody against Protein A and an ideal antibody against Protein B. The antibodies are...isotype? Antibodies are composed of light chains and heavy chains made up of variable regions and constant...interact. You read extensive research on the two proteins and come up with a great experimental plan that...detect Protein B and successfully visualize both proteins.     Figure 2: Two primary antibodies ...In most cases, the resulting chimeric antibody retains the binding specificity to the original target ...target and can be used with secondary antibodies against the new Fc region. In addition to increasing multiplexing...     Morelock, M. M., Rothlein, R., Bright, S. M., Robinson, M. K., Graham, E. T., Sabo, J. P., Owens, R.,...

  12. Four Ways to Package Transgenes That Exceed the Size Limit of Adeno-associated Virus

    Type
    Blog Post
    ...transgene.The AAV encoding part A of the transgene contains a promoter and the 5’-fragment of the transgene...donor. The AAV encoding part B of the transgene contains a splice site acceptor followed by the 3’-end ...methods. The AAV encoding part A of the transgene contains the promoter, the 5’ half of the transgene, a ...homology. The AAV encoding part B of the transgene contains the region of homology, a splice acceptor sequence...help prevent the accumulation of such truncated proteins. Examples of use:  Ghosh et al., 2008 used a...directed repairs (HDR) with CRISPR/Cas9. Donor A contains: 400bp homology arms to the site in the genome...stuffer DNA after the gRNA target site. Donor B contains: 400 bp homology arms to donor A,and “part B” ...

  13. March for Science

    Type
    Blog Post
    ...apolitical process for seeking knowledge. The process begins with a testable hypothesis - an educated guess ...scientists then revise their hypotheses and the cycle begins again. No part of this process is inherently political...States government? Politics and science are intrinsically linked: governments exist in reality and science...Environmental Protection Agency, and the National Institutes of Health endanger scientists’ ability to uphold...consequences While frustration and anxiety have inspired people to rally for science, others are concerned... themselves can come to conclusions themselves instead of depending on scientists or politicians to interpret...lines (18). Feeling like an apple will fall up instead of down when dropped will not make it happen. In...

  14. CRISPR 101: Homology Directed Repair

    Type
    Blog Post
    ... generally preferred for large insertions such as fluorescent proteins or selection cassettes. This class...conditional and insertion alleles using long ssDNA donors and CRISPR ribonucleoproteins. Quadros RM, et...Stroik January 24, 2023. DNA damage drives genome instability and contributes to cancer, premature aging, and...sticky or blunt ends and often introduces small insertions and deletions along the way. HDR uses homologous...overhang. This will serve as both a substrate for proteins required for strand invasion and a primer for ...template to create mutations HDR donors can contain insertable sequences or specific mutations, along with homologous...target sequence to be modified. In general, the insertion sites of the modification should be very close...

  15. Addgene-Seeding Labs plasmid grant round 3: microbial drug resistance and biofilms, antidiabetic plant components, visualizing brain injury - and a glowing robot fish

    Type
    Blog Post
    ...Plasmids expressing fluorescent proteins will be used to tag individual strains of bacteria. Their interactions...diabetes mellitus. Recombinant human proteins that play a role in insulin regulation and diabetes will be ...and the activity of the plant compounds on these proteins will be tested. Investigating changes in neuronal...after ischemic brain injury Dr. Gohar Tsakanova, Institute of Molecular Biology of the NationalAcademy of...condition is crucial. Plasmids expressing fluorescent proteins and a new two-photon laser scanning microscope...manipulate neural populations in order to get new insights into functional network interactions. Combining...bacterial strain. The sensor will be transported inside a small 3D printed robot that will “swim” in the...

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

    Type
    Blog Post
    ...Alex Chavez, researcher at the Wyss Institute for Biologically Inspired Engineering and member of the Addgene... vectors (~8.5 kb of insert) and the reduced titer associated with large inserts can make lentivirus production...particular, the Church lab at the Wyss Institute for Biologically Inspired Engineering and the Zhang and Doench.... For example, we’ve teamed up with the Broad Institute to provide lentiviruses for delivery of SpCas9...already expressing Cas9. As John Doench (Broad Institute and member of the Addgene Advisory Board) says...Doench/Root labs from the Broad Institute for providing us with advice and expertise as we’ve gotten these...

  17. Viral Vectors 101: Systemic Capsids

    Type
    Blog Post
    ...exists to prevent substances in the blood, like proteins and viruses, from crossing into the brain. While...increase the chances of inducing an immune response against the capsids or transgenes. Sensory neurons and ...as the type of transgene delivered: fluorescent proteins, optogenetic sensors, CRISPR elements, etc... ...capsids express differently in different mouse strains, a phenomenon that we are calling “strain-specific... but is more effective than AAV9 in many other strains, including FVB/NCrl, 129S1/SvImJ, DBA/2J, and C57BL...table from Huang et al, 2019 for affected mouse strains). However, other capsids cross the BBB via interactions...interactions with LY6C1 are effective in both mouse strains. It is therefore important to review the literature...

  18. 15 Years of Addgene: The Top 15 Plasmids

    Type
    Blog Post
    ... a VSV-G envelope expressing plasmid. Envelope proteins determine which types of cells a virus can infect...invariant sgRNA scaffold and cloning sites for inserting the guide sequence (pSpCas9(BB)). The 2A-GFP allows...pCMV-VSV-G was deposited by the Bob Weinberg lab and contains an envelope protein for producing lentiviral and...lentiviral packaging plasmid from the Trono lab contains the rev gene. It’s meant to be used with pMDLg...lentiviral packaging plasmid from the Trono lab that contains the gag and pol genes. It also requires pRSV-Rev...lab is a 3rd generation lentiviral vector that contains a scrambled shRNA. This scrambled shRNA is used... cloning vector. Tet-pLKO-puro - Tet-pLKO-puro contains all the necessary elements for packaging, reverse...

  19. Antibodies 101: Isotypes

    Type
    Blog Post
    .... Antibody heavy chain proteins as well as the genes that encode those proteins are designated by the ...antibody isotypes. Binding to complement system proteins, which can initiate a cascade that helps attack...their own. The antibody’s heavy-chain constant domains, or Fc region, can be bound by special receptors...location of disulfide bonds, the number of constant domains, and the hinge region length. Now for some essential...minor differences in their heavy chain constant domains. These are typically numbered, i.e. IgG1, IgG2,...IgG3, and IgG4. IgE IgE antibodies help protect against parasites but also play a role in allergies. They... any of the other isotype heavy chain constant domains. This switching helps B cells adapt during the ...

  20. Viral Vectors 101: Inverted Terminal Repeats

    Type
    Blog Post
    ...binding with Rep proteins and mutations in the trs site reduce Rep nicking. Since Rep proteins are important...with the viral rep (replication) and cap (capsid) proteins, which bind to the ITRs and initiate replication... with ITRs that flank the gene cassette, which contains a promoter, the gene of interest, and a polyA ...flip” or “flop” (Figure 2). Additionally, the ITR contains a 4-nucleotide Rep binding element (RBE) that ...resolution site that serves as the target site for Rep proteins (Savvy et al. 2013). Figure 2: AAV2 wild type...type ITR in flip and flop orientation. The ITR contains three palindromic sequences (A-A’, B-B’, and C-C...sequences. Using recombinant-deficient bacterial strains such as NEB Stable, Stbl3, JC8111, SURE2, or XL10...
Showing: 281 - 300 of 1050 results