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We narrowed to 317 results for: POR C

Showing: 181 - 200 of 317 results

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

    Type
    Blog Post
    Published
    June 4, 2024, 1:15 p.m.
    ...similar nuclease folds found in other proteins. The C-terminal domain (CTD) of the protein also belongs ...https://doi.org/10.1038/s41586-022-04470-1 Cofsky, J. C., Soczek, K. M., Knott, G. J., Nogales, E., & Doudna...Sternberg, S. H., Redding, S., Jinek, M., Greene, E. C., & Doudna, J. A. (2014). DNA interrogation by the...it is limited to only 20 bases, or why it’s so important to choose a target sequence with a nearby protospacer-adjacent... lobes laying open like a book (Figure 2). The portion of the CTD that recognizes the PAM is so flexible..., but they improve cutting efficiency and are important for robust activity in cells (Jiang & Doudna, ... guide RNA stabilizing the DNA-binding protein portion of the CTD, ready to check for PAM sequencines....

  2. Performing In Vivo CRISPR Screens Using the FITS Approach

    Type
    Blog Post
    Published
    May 2, 2024, 1:15 p.m.
    ...References and resources Milling, L. E., Markson, S. C., Tjokrosurjo, Q., Derosia, N. M., Streeter, I. S....screens have been used to interrogate T cells, an important immune cell type, in more complex and physiological...

  3. QC Sequencing Technologies at Addgene

    Type
    Blog Post
    Published
    July 15, 2025, 1:15 p.m.
    ...reactions, where a fraction of one of the four bases (A, C, T, or G) is supplied as a dideoxyribonucleoside triphosphate...regions of interest, or complex sequences such as G-C rich areas, secondary structures, or repetitive regions... technology, but Oxford Nanopore is one of the most popular options. Nanopore sequencing deviates from...pulled through the pore via motor proteins. When each nucleotide passes through the nanopore, the electrical...3: Overview of Nanopore sequencing technology. Created with BioRender.com. Nanopore can provide longer...sequencing methods (Figure 3). It uses (aptly named) “nanopores” that are attached to the flow cell membrane and...-read sequencing. At Addgene, we routinely use Nanopore technology to sequence our packaged viral DNA ...

  4. Proximity Labeling: A Powerful Tool for Protein Complex Purification and Proteomic Mapping

    Type
    Blog Post
    Published
    Dec. 2, 2019, 2 p.m.
    ...these methods a try.  References:  Branon, Tess C., et al. "Efficient proximity labeling in living cells...1837. PubMed PMID: 30676735. Kim, D. I., Jensen, S. C., Noble, K. A., KC, B., Roux, K. H., Motamedchaboki...the mitochondrial calcium uniporter (Martell et al., 2012). It is important to note that the biotin-phenol... of exogenous biotin allowing for more precise temporal control.  Find plasmids for TurboID and miniturbo... where two proteins are fused to complementary reporter proteins that can only assemble when in close ... ascorbate peroxidase as a genetically encoded reporter for electron microscopy." Nature biotechnology...proteomics approach to monitor the composition of spatiotemporally defined protein complexes." Nature communications...

  5. Hot Plasmids Spring 2024

    Type
    Blog Post
    Published
    May 23, 2024, 1:15 p.m.
    ...specificity of GFP expression for the indicated ssAAV5s. C) Quantifications showing high microglia transduction... microglia-targeting AAV plasmids here! Serrano, C., et al. (2023). Simple and Highly Specific Targeting...2024). They developed a new prime editor that incorporates La’s N-terminal domain, PE7 (Figure 2).  PE7...high microglia-specific gene expression when incorporated in an AAV vector (Figure 5). Efficiency and ...including in human retinal organoids and live mice. Importantly, REVeRT avoids the introduction of extraneous...associated with viral vector-based therapies. REVeRT supports advanced gene expression regulation, enhancing...

  6. The Fluorescent Vegetables in Aptamer Soup

    Type
    Blog Post
    Published
    Jan. 29, 2019, 2:27 p.m.
    ...due to poor folding and thermal instability at 37°C (Ouellet et al., 2016) and requires a tRNA scaffold...imaging single molecules and for live-cell imaging in C. elegans. Other groups have used Mango to study RNA-protein...split aptamer is composed of a “Top” and “Bottom” portion, each fused to a separate piece of RNA. Fluorescence...photostable, and were presented in the study to support the uniqueness of the photostability of the Corn-DFHO...growth as RNA biology, we are looking forward to reports on the next additions to aptamer soup! References...

  7. Tips and Tricks for Using Golden Gate Modular Cloning (MoClo)

    Type
    Blog Post
    Published
    July 8, 2025, 1:15 p.m.
    ...frozen glycerol stocks. It is very important that these stay frozen at -80 °C, since multiple rounds of freezing...; CDSns, coding sequence with no stop codon; TAG, C-terminal tag; TER, terminator. Created with BioRender.com... perfect Our first tip, and arguably the most important, is to take some time to sit down, plan your cloning...

  8. Getting the Most from Your Lentiviral Transduction

    Type
    Blog Post
    Published
    April 7, 2016, 2:30 p.m.
    ...into single-use aliquots, and stored at -80°C. Some reports suggest rapidly freezing virus in a dry ice... for less than a day, lentivirus can be kept at 4°C. For long-term storage, viral preps should be divided...protein attaches to cell surface receptors, other important binding events take place during the course of...

  9. Plasmids 101: E. coli Strains for Protein Expression

    Type
    Blog Post
    Published
    Feb. 10, 2015, 3:06 p.m.
    ...conditions. Many recombinant proteins express better at 30°C or room-temperature, which is accomplished by growing...growing your culture to the desired density at 37°C and reducing the temperature or moving it to a bench-top...added to the growth medium. Induction timing is important, as you typically want to make sure your cells... optimal because of the high cell-density they support; however, minimal media supplemented with M9 salts...

  10. Tips for CRISPR Gene Editing in Mice

    Type
    Blog Post
    Published
    June 28, 2016, 10:59 a.m.
    ...entire gene? Perhaps you are more interested in the c-terminus and would like to target that region. Or ... can be cloned directionally into the BbsI site. (c) The pCAG-EGxxFP target plasmid was co-transfected...expressing Cas9 and single guided RNA." Scientific reports 3 (2013). PubMed PMID: 24284873. PubMed Central...

  11. Delivery Methods for Generating iPSCs

    Type
    Blog Post
    Published
    April 17, 2018, 1:37 p.m.
    ...Zhi Ying Chen, Robert C. Robbins, Mark A. Kay, Michael T. Longaker, and Joseph C. Wu. "A nonviral minicircle...a Sendai Virus-Based Vector Using a Nonstructural C Protein and Endogenous MicroRNAs." Plos One 11, no...of Different Methods for Generating iPSCs. One important note: in this table, reprogramming efficiencies...high is based on the reprogramming efficiencies reported in González et al. A percentage is not listed ... be used to package reprogramming factors. An important milestone of reprogramming is the silencing of...cells for the presence of transgenes is still an important control. Additionally, Sendai virus doesn’t rely...comprised of OCT4, SOX2, LIN28, NANOG, and a GFP reporter. Minicircles are removed from cells by dilution...

  12. Targeting HIV-1 with CRISPR: Shock and Kill or Cut it Out?

    Type
    Blog Post
    Published
    May 10, 2016, 2:30 p.m.
    ...’s most common diseases. References Zhang Y, Yin C, Zhang T, Li F, Yang W, Kaminski R, Fagan PR, Putatunda...Gendron P, Zhu W, Guo F, Cen S, Wainberg MA, Liang C. CRISPR/Cas9-Derived Mutations Both Inhibit HIV-1 ...infection, including both common infections and opportunistic infections not usually seen in healthy individuals... genome had been excised, leaving only a small portion of the LTRs joined together. Clonal Cas9/gRNA-expressing...populations were also immune to HIV-1 reinfection. Importantly, no off-target effects were detected. Kaminski...patients. Although this is an exciting finding, it’s important to note that these HIV-1 patients were ART-naive...Cas9. While these results are discouraging, it’s important to note that these experiments were done in cell...

  13. Evolution of Brainbow: Using Cre-lox for Multicolor Labeling of Neurons

    Type
    Blog Post
    Published
    April 24, 2015, 2:39 p.m.
    ...Sato T, van Es JH, van den Born M, Kroon-Veenboer C, Barker N, Klein AM, van Rheenen J, Simons BD, Clevers...melanogaster. Hadjieconomou D, Rotkopf S, Alexandre C, Bell DM, Dickson BJ, Salecker I. Nature Methods. ...the 1980s, is one of the most important ways to spatially and temporally control gene expression, especially...ten colors (see table to the right); in various reports, Sanes and Lichtman have observed from 90-160 distinct...undergone recombination. This default state was disproportionately expressed by a majority of neurons, limiting... 8 colors. The use of AAV provides spatial and temporal control without the need for germline modification...refinements of the Brainbow technique will provide important insights into the complicated physical organization...

  14. Fluorescent Proteins 101: History of Fluorescent Proteins

    Type
    Blog Post
    Published
    Aug. 7, 2017, 1:58 p.m.
    ...expressed GFP in exogenous organisms (E. coli, and later C. elegans) for the first time. After Chalfie’s work...responsible for encoding GFP, and, in 1992, he reported its sequence. Soon after that, in 1994, Prasher...he lived! His hunch paid off, and he was able to report new fluorophores like DsRed from Discosoma sp. ...provided by the studies in Tsien lab, covered a portion of the color spectrum that GFP and its derivatives...reach. Nobel Prize for fluorescent proteins The importance of fluorescent proteins was recognized in 2008...

  15. The Importance of a Fun Workplace: Company Culture at Addgene

    Type
    Blog Post
    Published
    Sept. 3, 2015, 2:30 p.m.
    ...involve a huge group getting from point A to B to C to D, etc. 7. Don’t let a tiny [or non-existent] ...that company culture and enjoying your job were important parts of team buidling - they relieved some of...

  16. Antibodies 101: Single Chain Fragment Variables (scFvs)

    Type
    Blog Post
    Published
    June 3, 2021, 1:15 p.m.
    ...solubility (Monnier et al., 2013). The linker keeps the C-terminus of one variable domain and the N-terminus...10.3390/antib2020193  Khantasup K, Chantima W, Sangma C, Poomputsa K, Dharakul T (2015) Design and Generation...cells to make CAR T cells. The extracellular scFv portion of the receptors recognizes an antigen of interest...interest while the intracellular T cell receptor portion of the CAR facilitates signal transduction and release...

  18. Your Top Requested Plasmid in 2016!

    Type
    Blog Post
    Published
    Jan. 11, 2017, 8:47 p.m.
    ...site). Finally, in addition to the 2A peptide on its C-terminus, SpCas9 is also fused to a 3X FLAG tag at...are some additional features. The first and most imporant is that, upstream of the Cbh promoter, this plasmid...

  19. Plasmids 101: Gibson Assembly and Other Long-Homology Based Cloning Methods

    Type
    Blog Post
    Published
    March 1, 2016, 3:30 p.m.
    ...reaction takes an hour or less to complete at 50 °C. After an hour or so, the sample is immediately ready...PMID: 25861928. 3. Jiang W, Zhao X, Gabrieli T, Lou C, Ebenstein Y, Zhu TF. Cas9-Assisted Targeting of CHromosome...primers for PCR amplification of fragments to incorporate such regions of homology. This video gives a ...

  20. Management for Scientists: Seeking Feedback

    Type
    Blog Post
    Published
    Jan. 8, 2015, 2:34 p.m.
    ...ask at least one question to clarify the situation; C = confirm that you have understood the speaker accurately...focus on an aspect of communication especially important for a manager – getting feedback from the team... This can take practice and training. This is important because it creates a culture of engagement in ...  Seeking feedback frequently One of the most important paths of information flow is the one that brings...
Showing: 181 - 200 of 317 results