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  1. Plasmids 101: Using Transposons in the Lab

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    Blog Post
    ...somewhere else (Fig. 2). Class II TEs have LTRs on both of their ends.    Figure 2: Overview of DNA...has four major components: 1) a CRISPR RNA (crRNA), 2) four proteins (TniQ, Cas8, Cas7, Cas6) that forms... yeast, plant, insect, and mammalian cells, including human cells. piggyBac is biased to integrate at ...is active in vertebrates and integrates in human cells at rates similar to retroviral vectors. The hyperactive...piggyBac has an integration efficiency in human cells similar to retroviral vectors. The hyperactive PB...transposase (hyPB) is ~10-fold more active in mammalian cells compared to the codon-optimized wild-type piggyBac...G/C)TNA. Tol2 can deliver 10-11 kB to mammalian cells without decreased efficiency, with a maximum cargo...
  2. Antibodies 101: Conventional vs Spectral Flow Cytometry

    Type
    Blog Post
    ...from another within a fully stained sample. Figure 2 is an example of the emission spectra for two fluorophores...another during spectral unmixing.     Figure 2: The full emission spectra for BV421 and BV711 on ...co-expression among cellular markers You’ll also need to know if markers are co-expressed in your cells when designing...immunologists, allowing for the rapid analysis of cell populations within a heterogenous tissue type, such...methods, the identification and isolation of rare cell types. In recent years, full spectrum flow cytometry...detector to detect one fluorophore and therefore one cellular marker. However, emission spectra of certain fluorophores...Spectral flow allows for autofluorescence extraction Cells produce a low level of fluorescence called autofluorescence...
  3. Simple CRISPR-based Epigenetic Editing: dCas9-directed DNA Demethylation

    Type
    Blog Post
    ...targeted DNA demethylation. Cell Discov 2, 16009. https://doi.org/10.1038/celldisc.2016.9.   ...Hrit et al. 2018, Kaas et al. 2013). Figure 2: CRISPR TET-based epigenetic editing. In reality, ...its oxidized derivatives. Cell 152, 5 P1146-1159. https://doi.org/10.1016/j.cell.2013.02.004.  Xu X, Tao...enzymatic activity cause demethylation?  Dividing cells depend on DNA methyltransferase 1 (DNMT1) to copy...methylate sites blocked by dCas9. Within a few rounds of cell division, the original methylated strand is diluted...dCas9 expression and pLM-CMV-R-Cre. Selection for cells stably and highly expressing these constructs can...can be achieved by fluorescence-activated cell sorting, and stable gRNA expression can be achieved using...
  4. Antibodies 101: Antibody Engineering and Directed Evolution

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    Blog Post
    ... follow the same general cyclical process (Figure 2). The differences typically come in when generating...starting point for the next cycle. Figure 2: General process of antibody directed evolution. Created...module_attribute "schema_version" is_json="true" %}{% raw %}2{% endraw %}{% end_module_attribute %}{% module_attribute...high-throughput screening techniques such as ELISA and cell surface display are common. ELISA enables quantitative...for comparing relative affinities across mutants. Cell surface display systems, such as bacterial, yeast...presentation of your mutant libraries on the surface of cells. This enables direct screening via flow cytometry...provide recombinant antibodies that target families of cell surface and secreted proteins (glypicans, integrins...
  5. Plasmid-based Recombinant Monoclonal Antibodies: What They Are and Why You Should Be Excited About Them

    Type
    Blog Post
    ...from animals or hybridomas.   Figure 2: (Left) Figure 2: (Left) The light chain and heavy chain genes... be introduced into host cells, such as bacterial, yeast, or mammalian cells, for antibody production ...Nanobodies and recombinant binders in cell biology. Journal of Cell Biology 209:633–644 . https://doi.org...animals and bleed dates and even clonal hybridoma cell lines can produce more than one mAb (Bradbury et...plasmid or from an integrated sequence in a stable cell line. The genes encode the heavy chain and light...antibody protein binds an antigen. Image from Fvasconcellos.     How are Recombinant Antibodies Made...
  6. Illuminating Epigenetics with A FRET Based Biosensor

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    Blog Post
    ...FRET signal back again. The Ting lab constructed 2 versions of this reporter to target the K9 (pcDNA3... 525.7570 (2015): 533-537. PubMed PMID: 26352475. 2. Bygren, Lars O., et al. "Change in paternal grandmothers...genes should be ‘read’ by the cellular machinery. This is how most cells in the body contain the same ...complex of DNA and histone proteins that compacts cellular DNA into chromosomes and reduces the risk of dangerous...the more compacted it becomes. This prevents the cellular machinery from physically interacting with the...visualise changes in histone methylation in living cells. Essentially this reporter construct consists of...recognise lysine 27 methylation. When tested in living cells where methylation of these sites was controlled,...
  7. Plasmids 101: Terminators and PolyA signals

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    Blog Post
    ...due to the presence of additional helper sequences (2-3). As alluded to above, termination and polyadenylation...and translation processes as well.   Figure 2: The conserved eukaryotic polyadenylation signal directs...Plasmids designed to express genes in a given host cell type are generally broken down into two broad categories...purpose and mechanism of polyadenylation vary across cell types, but polyadenylation generally serves to promote... thought that poly(A)s are used to control the cellular concentration of regulatory RNAs and may additionally...additionally act as a quality control mechanism to rid the cell of mis-folded RNAs. Eukaryotic termination and polyadenylation...signal by using in vitro analyses. Molecular and Cellular Biology. 1992. PMID: 1333042. PubMed Central PMCID...
  8. 7 Tips to Secure a STEAM Internship This Summer

    Type
    Blog Post
    ...and scholarships in different subject areas!    2. Resume and cover letter know-how  Tailor your resume...4. Letters of recommendation wisdom  Reach out to 2-3 individuals in your professional circle to vouch...college. Prior to March 2020, internships were a fiercely competitive endeavor. Due to the COVID-19 pandemic...
  9. Form Your Own Peer Mentoring Group: A How-To Guide for Scientists

    Type
    Blog Post
    ...The groups in these programs typically consist of 1-2 mentors and 3-5 mentees who meet together for an 8... together. If you all work at the same place, a 1-2 hour lunch on a regular schedule might work. If not...do once you are in a mentoring relationship.  Celebrate your successes and support each other as you transition...
  10. 28 Hot Plasmid Technologies from 2015

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    Blog Post
    ... Zetsche et al., Nat Biotechnol. 2015 Feb 2;33(2):139-42. doi: 10.1038/nbt.3149. Light-inducible ...and on your favourite signalling pathways (Figure 2). They are all available now at Addgene. 1 Lungu...20;19(4):507-17. 10.1016/j.chembiol.2012.02.006. 2 Guntas et al., Proc Natl Acad Sci USA 2015 Jan 6; ...Nihongaki et al., Chemistry and Biology. 2015 Feb 19; 22(2):169-74. High efficiency cloning for enzyme screening...inducible, gene knockdown with shRNA or shRNA-miR30, (2) tamoxifen-inducible CreERT2 gene deletion, (3) simultaneous...different fluorescent proteins, 8 constitutive promoters, 2 includible promoters, 3 polyA terminators and various...of plasmids required for each input/output system; 2) removing all chemically inducible promoters; 3) improving...
  11. Adeno-associated virus (AAV) Guide

    Type
    Guide
    ...capsids from serotypes 1, 2, and 5 display differential efficiency and cell tropism after delivery to ...packaging and producer cell lines for the production of AAV vectors . Microorganisms, 12 (2), 384. https://doi.org...components, see the AAV Plasmid Elements table . Figure 2: Plasmids required for rAAV vector production. Created...the central nervous system . Molecular Therapy, 10 (2), 302–317. https://doi.org/10.1016/j.ymthe.2004.05.024...retrograde access to projection neurons . Neuron, 92 (2), 372–382. https://doi.org/10.1016/j.neuron.2016.09.021...based on other human cell lines, such as HeLa and A549 cells, and even insect Sf9 cells , have also been ...able to infect cells and replicate autonomously. Human embryonic kidney 293T (HEK293T) cells are the most...
  12. Retrograde AAV viral preps

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    Collection
    ...Calcium sensor Ting 83899 pAAV-mDlx-GCaMP6f-Fishell-2 mDlx GCaMP6f expression under the control of the mDlx... of Cre, and expresses NLS-EGFP in Cre-positive cells. Control Harvey 137164 pAAV-nEF-Con/Fon/Von eYFP...
  13. 5 Tips for Troubleshooting Viral Transductions

    Type
    Blog Post
    ...Central PMCID: PMC3760253. 2. Rohwer, Forest. "Global phage diversity." Cell 113.2 (2003): 141. PubMed ... digest and compare it to some reliable controls. 2. Viral stocks may be sensitive to freeze thaw cycles...contaminating your transduction with the packaging cells. Packaging cell debris can be removed either by filtration... you can reduce the volume of cell culture medium on the packaging cells immediately following transfection...efficiency in cells that are sensitive to the toxicity of polybrene (e.g., hematopoietic or primary cells), and... the packaging cells after 1–3 days of selection. A lower percentage of surviving cells may be the result...adeno-associated virus transduction of cells." American Journal of Physiology-Cell Physiology 291.2 (2006): C386...
  14. Viral Vectors 101: Voltage Indicators

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    Blog Post
    ...Indicators GEVIs can be divided up structurally into 2 main groups: voltage sensitive domains and Rhodopsin...depending on your experimental system.   Figure 2: General overview of chemigenetic tools. A genetically...Cellular communication and function depend on potential energy. Cellular activities like action potentials...Indicators As previously mentioned, cellular voltage is measured at the cell membrane, and this voltage is ...endocrine cells. It follows that the specific targeting of these voltage indicators needs to be cell type ...electrical events! Voltage indicators are voltmeters for cells! In this blog we will review how these indicators...related to the electrical impulse of the cardiac cells. To study this phenomenon you will likely want to...
  15. Fluorescent Proteins 101: Green Fluorescent Protein (GFP)

    Type
    Blog Post
    ...from GFP and the mutation definitions found in Table 2 may not apply. Table 1: The specific mutations comprising..., V163A EBFP F64L, S65T, Y66H, Y145F   Table 2: Functional role of specific mutations in GFP derivatives...measure, and control molecular signals, cells, and groups of cells in order to understand their activity...Fluorescence-activated cell sorting (FACS): This is a type of flow cytometry that separates mixtures of cells into distinct...Thus, FACS can be used to separate cells expressing GFP from cells that are not. Developmental/transgenic... localization profiles, acting as a biosensor or cell marker, studying protein-protein interactions, visualizing... was non-toxic and could be expressed in living cells, enabling the study of dynamic, physiological processes...
  16. Deep Dive: Fixing and Permeabilizing for Immunofluorescence

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    Blog Post
    ...membrane integrity (Fig. 2). For example, saponin interacts with cholesterol in cell membranes and leaves ... at too high a concentration.      Figure 2: Examples of different permeabilization strategies...holes in the cell membranes of your sample to allow antibodies better access to intracellular targets. If...allowing antibodies into the cell where they could bind to intracellular epitopes. C) Permeabilizing with...method for visualizing molecules of interest within a cell or tissue. In that introduction, we broke down the...structure of your samples, the relationships between cellular components, and the accessibility of your target...cross-linkers work by (as the name suggests) cross-linking cellular proteins. Cross-linking is a chemical reaction...
  17. Plan Your Experiment

    Type
    Guide
    ...your target cells. There are a variety of ways to deliver and express these components (Figure 2), and the...optimal system will likely depend on your cell type. Figure 2: Different delivery systems utilized for ...and/or Cas9 expression or (2) a lack of efficient target cleavage in cells that do express both Cas9 and...CRISPR-Based genome editing. Trends in Biotechnology , 36 (2), 173–185. https://doi.org/10.1016/j.tibtech.2017.11.006...difficult-to-transfect cells. You can direct expression to specific cell types by using cell-specific promoters...human embryonic kidney 293 (HEK293) cells. For more difficult cell types, you may need to look into other...are good choices for more difficult cell types like primary cells. For CRISPR delivery, lentiviral and...
  18. Google Forums Round Up: First Impressions of NgAgo

    Type
    Blog Post
    ... PMID: 27136078. 2. Burgess, Shawn., et al. "Questions about NgAgo." Proteins & Cell (2016). 3. "NgAgo...engineering methods have their own pros and cons (2). The NgAgo DNA guide is inexpensive and can be 5’...degraded by the cellular machinery, leaving a small amount of active NgAgo/ssDNA complex in the cell. As proposed...results of Gao et al 2016 in a letter to Protein & Cell. Update (August 3rd, 2017) THE ORIGINAL NgAgo ARTICLE... guide, it cannot be produced from a plasmid in cells. Additionally, in vitro assembly of NgAgo/ssDNA ...dangerous, non-physiological temperature for mammalian cells and one which diminishes the endonuclease activity...guide. Therefore, researchers must co-transfect cells with 5’-P-ssDNA guides and an NgAgo expression plasmid...
  19. Chromoproteins: Colorful Proteins For Molecular Biology Experiments

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    Blog Post
    ...amilCP, tsPurple, and gfasPurple) (Figure 2). Figure 2: Chromoprotein expression from bacteria on... them into your cells, you can simply look for blue colonies to determine which cells have taken up the... that the cellular fitness cost varied greatly between chromoproteins. In some cases, cells containing... mScarlet as the donor in HeLa cells.   Find ShadowR for mammalian cell expression here   Now that you...cloning experiment worked just by looking at the cells on the plate. For example, if your plasmid contains...Here, the chromoprotein is expressed and gives the cell a magenta (amilCP) or purple (tsPurple) color. Though...added which induces expression of the toxin. Any cells that grow after this point will be those that have...
  20. Empty Backbones - Choosing Your Perfect Plasmid Backbone

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    Collection
    ...backbone with MS2 loops at tetraloop and stemloop 2 and EF1a-zeo resistance marker pLenti CMV/TO GFP-Zeo...and display of proteins on the extracellular surface of S. cerevisiae cells His-tagged versions of pFastBac... the elements needed for replication in the host cell and are designed for a user to insert a gene of ..., or express the insert they carry in the target cell. These vectors allow you to test the function of....1-mCherry - Expresses mCherry tag in mammalian cells pBI-MCS-EGFP - Tet-inducible Find more Tet-inducible...or Fusion Protein Tags and fusion proteins are excellent tools for further understanding the function... as GFP, which allows you to visualize the cellular localization of your protein. Additionally...
  21. Pooled Library Amplifications

    Type
    Blog Post
    ...estimate transformation efficiency (Figure 2).    Figure 2: Example of a serial dilution to estimate...Genome-wide Inhibition Library. DNA and cell amounts After deciding on a cell type, you will need to determine...DNA and how many cells to use for your transformation(s). The amount of competent cells you need total ... generally electroporated into electrocompetent cells to ensure the highest possible transformation efficiency...efficiency. Using Stbl4 or other ultra-high efficiency cells ensures that plasmid diversity in the library is...transformation may vary depending on the particular type of cells you are using, as well as other factors, such as...transformations, such as splitting 400 ng of DNA in 100 μL of cells over four individual electroporations. The specifics...
  22. Antibody Guide

    Type
    Guide
    ...surface of most immature B cells; function is unknown Monomer with a valency of 2 IgE Protects against parasites...such as anaphylactic shock Monomer with a valency of 2 IgG The most abundant isotype in blood, also found...secondary immune response Monomer with a valency of 2 IgG1; IgG2; IgG3; IgG4 IgM Produced during the primary... ranging from antibody fragments (shown in Figure 2) to completely unrelated molecules. Examples include...used for assembling protein nanostructures. Figure 2: An antibody (IgG, far left) compared to alternative... produced. Cells - Antibodies can also be produced in cell cultures using hybridomas, cell cultures created...created by fusing B cells with myeloma cells (immortalized B cells). The genetic reorganization that creates...
  23. CRISPR Activators: A Comparison Between dCas9-VP64, SAM, SunTag, VPR, and More!

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    Blog Post
    ...gene activation, with some genes experiencing around 2-fold activation levels. Although other methods have.... This method has been shown to activate genes at 2- to 3-fold higher than SAM, SunTag, and VPR (Zhou ...Control of Gene Expression. Cell 152:1173–1183 . https://doi.org/10.1016/j.cell.2013.02.022 Sajwan S, Mannervik...and Fluorescence Imaging. Cell 159:635–646 . https://doi.org/10.1016/j.cell.2014.09.039 Zhou H, Liu J,...from when looking to activate genes across diverse cell lines.   dCas9-VP64 Description CRISPR activation...large and are not expressed consistently throughout cells. VPR Description VPR fuses a tripartite complex ...activation levels than SAM in some loci across Drosophila cell lines (Sajwan et al., 2019). SPH SPH combines components...
  24. 6 Tips for Grant Writing

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    Blog Post
    ...days before the deadline isn’t going to be much use. 2. Originality is essential for grant success The two... it’s important not to throw in the towel if 1 or 2 funding bodies say no. If possible, get feedback from...research (either because of expertise, equipment, or excellence of the institution). Likewise, if you’re staying...Cambridge University where he studied mechanisms of cell division. Recently Seán got the entrepreneurial ...
  25. Twenty Years of Sharing Science!

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    Blog Post
    ... anniversary! We've done so much in the past two (2!) decades that it's hard to summarize, but we wanted...into the Addgene website search bar?   We'll be celebrating all year long, so keep an eye on our blog and...
  26. PITChing MMEJ as an Alternative Route for Gene Editing

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    Blog Post
    ... PITCh vector via In-Fusion or SLIC cloning. Step 2: Design an insertion locus-specific gRNA The gRNA ...Find the plasmids from this publication at Addgene. 2. Dalvai, Mathieu, et al. (2015). “A Scalable Genome-Editing... for Mapping Multiprotein Complexes in Human Cells.” Cell Rep. 13: 621-633. PMID: 26456817. Find plasmids...Chromosomes). Using their PITCh plasmids, GFP knock-in cell lines can be created in about a month and a half...locus-specific gRNAs Step 4: Select for puromycin resistant cells Step 5: PCR amplify and sequence the locus to verify...crispr.mit.edu. They tested their system in HEK293 cells, integrating the GFP-Puro cassette into the FBL ...
  27. Plasmids 101: Degron Tags

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    Blog Post
    ...required components are 1) a SMASh-tagged protein and 2) asunaprevir (small molecule). dTAG The degradation...gone through an additional round of evolution (AID-2) to reduce some of the leakiness associated with the...uses a small molecule, a tag, and the endogenous cellular degradation machinery to function. The dTAG is...tag and small molecule. It does work in mammalian cells, mice, and even yeast, though (we have a kit!). ...and complete degradation of proteins in mammalian cells. Proc Natl Acad Sci U S A. 2012 Dec 4;109(49):E3350...Tags Plasmids for Endogenous Gene Tagging in Human Cells Antibodies 101: Epitope Tags  ...
  28. With an Eye Towards the Future, We Look Back at the March for Science

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    Blog Post
    ...Vanderbilt University. She studies beta-cell function and failure in Type 2 Diabetes. Birmingham, AL, USA 33.5207...scientists have 1) the courage to make mistakes and 2) the integrity to recognize these mistakes quickly...available to people who want to join the dialog and 2) remember that science should not be partisan: everyone...of high profile people denying science, from celebrities falsely claiming vaccines cause autism to politicians...worried the march would be a simple, unquestioning celebration of science, at a time when scientists must rise...BC (Canada). She works in a lab that uses yeast cells to study how DNA damage promotes cancer. She is ...
  29. Better Dyeing Through Chemistry & Small Molecule Fluorophores

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    Blog Post
    ...26000849. 2. Grimm, Jonathan B., et al. "A general method to improve fluorophores for live-cell and single-molecule...expressed as a fusion with your favorite protein and (2) a synthetic fluorophore-contianing “ligand” that ...). These cell-permeable Janelia Fluor (JF) dyes have excellent properties inside living cells, especially... fluorescent proteins. In-cell labeling strategies The majority of in-cell labeling strategies have two...and oligonucleotide labels for fixed-cell imaging, not for live-cell applications. Thus, these fluorophores..."Imaging live-cell dynamics and structure at the single-molecule level." Molecular cell 58.4 (2015): 644...sparked a renaissance in biological imaging. Suddenly, cell biologists were no longer beholden to chemists and...
  30. Plasmids 101: Choosing an Antibiotic Resistance Gene

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    Blog Post
    ...suppress Zeocin-induced DNA cleavage in human cells. Mutagenesis, 20(2), 111–114. https://doi.org/10.1093/mutage...antibiotic selection. Essentially, this gene is a 2-in-1 providing both bacterial and eukaryotic selection...prevent cell death. One of the cool things about amp is that it's exclusively toxic to dividing cells. The... continuous cellular processes (e.g., protein synthesis), while amp only affects the cell wall synthesis...Check icon" %} Spectinomycin Excellent No Just bacteria Not SHuffle cells {% icon icon_set="fontawesome...icon" %} Zeocin Excellent No Bacteria, plants, and eukaryotes Not Tn5-containing cells {% icon icon_set..., penicillin, and is a beta-lactam that inhibits cell wall synthesis. The AmpR (amp resistance) gene produces...
  31. Fluorescent Proteins 101: Fluorescent Protein Timers

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    Blog Post
    ...linear region to solve for total expression time. 2. Timer functionality with a single gene Although it...290.5496 (2000): 1585-1588. PubMed PMID: 11090358. 2. Tsuboi, Takashi, et al. "Age-dependent preferential...to monitor cell, organelle, and protein localization. For instance, you might dye your cells and look at...either in whole cells or as a pool of fusion proteins localized to a specific region of the cell. For example...the relationship between the age of a cell, protein, or cellular structure and a particular biological...trafficking to a subcellular location, start of gene expression, development of a cell structure, etc)....timers, researchers can look at a single image of a cell and understand how protein localization changes ...
  32. Antibodies 101: Producing Recombinant Antibodies

    Type
    Blog Post
    ...detection, and control. Int J Pharm. 2018 Oct 25;550(1-2):251-268. doi: 10.1016/j.ijpharm.2018.08.043. Epub...and particle formation. J Pharm Sci. 2015 Feb;104(2):602-11. doi: 10.1002/jps.24259. Epub 2014 Nov 21....and yeast cells are cost-effective and faster-growing than mammalian cells, but mammalian cells can perform...your cells with your plasmid(s), and then wait 7-14 days (in mammalian cells) to collect the cell culture...production system, cell lines typically used include Human Embryonic Kidney (HEK) cells and Chinese Hamster...recombinant antibodies with some molecular biology and cell culture experience. Let’s go over the basics of ... place in bacterial, yeast, plant, or mammalian cells. Each of these systems has their advantages and ...
  33. New Tools Enable CRISPRa for Neuroscience Applications

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    Blog Post
    ...with neuron-specific expression of dCas9-VPR (Figure 2), a popular Cas9 activator developed by the Church...ubiquitous ones such as EF-1α and CAG. Figure 2: Dual lentivirus system with ubiquitous expression...fluorescence activated cell sorting (FACS) to isolate differentiated from undifferentiated cells at the end of ...these cell types are used. The lack of good flow cytometric antibodies for many CNS-resident cell types...that drive neuronal fate and reprogramming." Cell stem cell 23.5 (2018): 758-771. PubMed PMID: 30318302...CRISPRa screening approach in embryonic stem (ES) cells to determine which transcription factors promote...antibodies, downstream of the TUJ1 coding sequence. ES cells that stably express the dCas9-Suntag activator were...
  34. Tips for Screening with Yeast Two Hybrid Systems

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    Blog Post
    ...retain their individual activities when separated (2), and that DBD and TAs from different systems could... Enzymol. 1983;100:293-308. PubMed PMID: 6312261. 2. Keegan, L., Gill, G., and Ptashne, M. Separation ...entire proteome of a particular cell type, or a subset based on a cellular compartment or growth stage)....extensively to screen for molecular interactions in the cell, including protein-protein, protein-DNA and protein-RNA...field of eukaryotic transcriptional activation and cell biology. During this period, proteins were successfully...The reporter gene then allows detection of those cells where binding between the proteins of interest has...alternative fungi (C. albicans, pC2HB) (7) and mammalian cells. If the problem is that your proteins are not post-translationally...
  35. Getting to Know Expansion Microscopy

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    Blog Post
    ...maybe label) your sample (i.e. cultured cells or tissues).  Step 2: attach anchors to your sample that will...quite so much. Created with biorender.com Steps 2-4 are not found in standard IHC/ICC protocols and ... for EM is not something you find in the average cell and molecular biology lab, and EM is limited in ...ultrastructural relationships that influence everything from cell division to memory, but don’t have access to the...properties of the sample itself. For example, cultured cells are typically easier to expand than mouse brain ..., Mohar B, Eddison M, et al (2022) Visualizing cellular and tissue ultrastructure using Ten-fold Robust... Zwettler FU, Guennec ML, et al (2019) Imaging cellular ultrastructures using expansion microscopy (U-...
  36. An Interview with Connie Cepko - Gene Therapy, Plasmid Tools and Insights to Success

    Type
    Blog Post
    ...from this publication at Addgene. 2. Tang, Jonathan C.Y., et al. “Cell type-specific manipulation with ...fluorescent proteins as scaffolds for cell-specific gene manipulation.” Cell 154(4) (2013): 928-939. PubMed ...Cepko Interested in Working on the Eye 9:05-10:16 : Cell Targeting in Retinal Gene Therapies 10:17-14:34 ...28:39 : Methods for Delivering Multiple Genes to Cells for Gene Therapy 28:40-35:10 : GFP Activatable Tools...
  37. Antibiotic Resistance: An Old Solution but a New Problem

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    Blog Post
    ...diffusion test or Kirby-Bauer susceptibility test (Figure 2), and requires at least 18-24 hours to obtain results...." London: Wellcome Trust & HM Government (2016). 2. CDC Information Pages on Antibiotics 3. WHO Antimicrobial... attack the cell wall of bacterial cells; however, Mycoplasma species do not possess cell walls. As a ...bacteriophage (viruses that only infect bacterial cells), or a bacterium picking up resistance genes from...conserved, non-negotiable process within the bacterial cell, antibiotic resistance is very likely to develop...
  38. CRISPR 101: Cytosine and Adenine Base Editors

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    Blog Post
    ... a template, copying the base edit. Figure 2: Cytidine deamination takes place on the free strand...SpCas9 base editors with editing windows as small as 1-2 nucleotides. To reduce off-target effects associated...formation. To further decrease indel formation 1.5-2 fold, the team fused bacteriophage protein Gam to ...which we've discussed in two follow-up posts: (1) (2). Base editors have a narrow editing window of only...that it appears “newly synthesized” to the cell. Thus, the cell repairs the DNA using the U-containing strand..., which may lead to cell death rather than NHEJ repair, thus removing these cells from the edited population...target modification in primary T cells, making them a promising tool for cell therapy applications. Dual base...
  39. Antibodies 101: Choosing the Right Isotype

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    Blog Post
    ... reduce the background staining (Table 2).      Table 2: Secondary antibody selections for a four-antibody...for Light and Electron Microscopy. In Methods in Cell Biology (pp. 283–310). Elsevier. https://doi.org...
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