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We narrowed to 586 results for: cel.2;

Showing: 161 - 170 of 586 results

  1. Controlling for Off-target Effects with a New Genome-wide CRISPR Screen Design

    Type
    Blog Post
    ...guide expression and dsDNA breaks. Figure 2: Non-targeting vs. safe-targeting guides. The genome-wide...target all protein-coding genes present in the genome; 2) non-targeting guides (red), which target no sequence...guides that had zero mismatch off-targets. Guides with 2-bp mismatch off-targets were only toxic if they had...tolerated than mismatches more distal to the PAM, and 2) guides with high GC content have greater off-target...PMID: 28474669. Pubmed Central PMCID: PMC5424143. 2. Tsai, Shengdar Q., et al. "GUIDE-seq enables genome-wide...delivered as a pooled library via a lentiviral vector to cell lines that either stably expressed Cas9 or that ...almost all genes involved in the production of a cell surface glycan that’s required for ricin uptake....

  2. 15 Hot Plasmids from 2017

    Type
    Blog Post
    ...NGG)  Editing window ~2 nt  85175  pBK-EE-BE3  SpCas9 (NGG)  Editing window ~2 nt  85176  pBK-YE2-...quantum yield of 0.54) and mScarlet-H with M164H (2-fold improvement in photostability compared to mScarlet...SpCas9 base editors with editing windows as small as 1-2 nucleotides! To reduce off-target effects associated...pBK-YE2-BE3  SpCas9 (NGG)  Editing window ~2 nt  85177  pBK-YEE-BE3  SpCas9 (NGG)  Lower on-target activity...activity than BE3; editing window ~1-2 nt  87438  pET42b-HF-BE3  HF-Cas9 (NGG)  Very low off-target activity...Ladders Are Awesome Tools: They are comprised of 2 plasmids, pPSU1 and pPSU2, that can be amplified affordably...libraries The Gross Lab at UCSF recently deposited 2 Bacillus subtilis libraries with Addgene – the B. ...

  3. Viral Vectors 101: Integrase-Deficient Lentiviral Vectors

    Type
    Blog Post
    ...or non-homologous end joining (see Figure 2).  Figure 2: Schematic representation of integration...developing T-cell acute lymphoblastic leukemia due to insertional mutagenesis (Howe et al., 2008). 2. Transient...vectors: progress and applications. Gene Therapy, 17(2), 150–157. https://doi.org/10.1038/gt.2009.135. PubMed...efficiently during infection. Journal of Virology, 70(2), 721–728. https://doi.org/10.1128/JVI.70.2.721-728.1996...purposes, like creating stable cell lines, delivering CRISPR/Cas9 to cells for genome editing, and introducing... chromosome of the cell. This integrated viral genome is then part of the cellular DNA… forever!  This...viral entry into the host cell.  Transfecting these three plasmids into cells enables the production of...

  4. Viral Vectors 101: What is a Virus (and a Viral Vector)?

    Type
    Blog Post
    ...developed around how they replicate (Figure 2). Figure 2: Baltimore replication classes. Image credit...safer viral vectors that can be used in BSL-1 or BSL-2 labs. Viral vectors are viruses that have been genetically...hijacking the machinery of the cells they infect. Once they enter a cell, they treat their viral genome...genome as a “payload” that is delivered to the cell. The cell then uses its own machinery to replicate the...replication process whereby one cell copies itself and “splits” into two cells. Viral vectors Viruses are ...virus. From one, come many After a virion infects a cell, the single particle creates many copies of itself...millions of new copies, which are released from the cell like baby spiders from an egg sac. (Okay, some enveloped...

  5. Technical Design of a Western Blot

    Type
    Blog Post
    .... The most common options are 4 °C overnight or 1–2 hours at room temperature (RT). Most people choose...overnight at 4 °C and their secondary antibody for 1–2 hrs at room temperature, which allows them to reduce...Tris-glycine 6–400 kDa Tris-glycine 100 V, 1–2 hours Easy and cheap to handcast Short shelf-life...–40 kDa Tris-tricine 30 V, 1 hour or 100 V, 1–2 hours Good separation, quality, and stability ...separation of proteins of different sizes.  Table 2: Recommended gel percentages for various protein sizes...are consistent and stable at room temperature for 1–2 years and are available in formulas specific to your...response over a broad dynamic range that typically spans 2–5 orders of magnitude, which includes a wide dynamic...

  6. Delivery Methods for Generating iPSCs

    Type
    Blog Post
    ... and future challenges." Journal of Cellular Physiology 228, no. 2 (2012): 267-75. PubMed PMID: 22767332...Pluripotent Stem Cells, 2013, 23-33. PubMed PMID: 23546745. PubMed Central PMCID: 4176696. 2. Yusa, Kosuke...Gene-delivery systems for iPS cell generation. Expert opinion on biological therapy, 10 2, 231-42. PubMed PMID:...plasmid-based expression requires serial transfection of 1 or 2 plasmids that express the reprogramming factors of...for generating iPSCs by plasmid transfection where 2 of 11 clones tested had plasmid integration. Another...understanding the mechanisms of reprogramming, and 2) studies with clinical endpoints. In the first scenario...PubMed PMID: 27764162. PubMed Central PMCID: 5072705. 2. Sommer, Cesar A., and Gustavo Mostoslavsky. "The ...

  7. A Needle in a Base-Stack: Cas9 Structural Biology

    Type
    Blog Post
    ...Cas9 from Streptococcus pyogenes.   Figure 2:  Crystal structure of S. pyogenes Cas9 in the apo...NUC and REC lobes laying open like a book (Figure 2). The portion of the CTD that recognizes the PAM is...it doesn’t appear in the crystal structure (Figure 2) at all. When this crucial recognition region is disordered...regions, so crystal structures like those in Figures 2 and 3 don’t show these regions, even if they are present...itself at the bottom of the image. Stem loops 1 and 2 extend horizontally across both lobes. The guide RNA...is required for Cas9 nuclease function. Stem loops 2 and 3 are not strictly necessary for in vitro cleavage...open conformation, but highly flexible, as in Figure 2. The guide RNA is bound primarily by the REC lobe,...

  8. Plasmids 101: Restriction Cloning

    Type
    Blog Post
    ... used. Follow the manufacturer’s instructions.   2. Isolate Your Insert and Vector by Gel Purification...DNA concentration alone. One method is to conduct 2 ligations for each plasmid you are trying to create...containing the ligation mixture with backbone alone; 2. Transformation containing ligation mixture with insert...colonies than the vector + insert plate (unsuccessful 2), or roughly equivalent numbers of colonies on each... or equivalent to backbone + insert, unsuccessful 2 and 3 above), you can try ligating the recipient plasmid...use electro-competent cells instead of the more common chemically-competent cells. You should perform, ... manufacturer’s instructions for your competent cells. For most standard cloning, you can transform 1-...

  9. No Llamas Required - Synthetic Nanobodies Against Membrane Proteins

    Type
    Blog Post
    ...select for optimal sybody candidates. Figure 2: Outline of the process for selection of sybodies ...identify and isolate sybodies targeting the SARS-CoV-2 Receptor Binding Domain found on the membrane-bound...Generation Toolbox kit. Find the Seeger lab's SARS-CoV-2 sybodies! References Geertsma ER (2013) FX Cloning...2020) Synthetic nanobodies targeting the SARS-CoV-2 receptor-binding domain. BioRxiv.  https://doi.org...Single-Domain Camelid Antibodies. Cell 181:1004–1015.e15 . https://doi.org/10.1016/j.cell.2020.04.031 Zimmermann ...coronaviruses and enable the virus to enter host cells. To do this, they injected the spike protein from...camelid antibodies that bind to the spike proteins in cell culture, and inhibit these viruses.  But llama ...

  10. Year of the Camelid: Antibody Style

    Type
    Blog Post
    ...and a source of sdAbs (Figure 2) (Xu, et al., 2021).      Figure 2: Genetically modified mice offer... from camelid mice and llamas neutralize SARS-CoV-2 variants. Nature 595:278–282. https://doi.org/10.1038...antigens in living cells or tissues. sdAbs can also be used to manipulate cellular functions, like in ...sdAbs over conventional antibodies include:  Excellent tissue penetration, thanks to their small size...
Showing: 161 - 170 of 586 results