We narrowed to 595 results for: cel.2

...Nutr, 129 (2009a): 477–484. PubMed PMID: 10064313. 2. Chapman-Smith, A., & Cronan, J. E. “The enzymatic...carboxyl groups to organic acids to produce important cellular metabolites. Biotin acts as a carboxyl carrier...chemically. Enzymatic biotinylation occurs naturally in cells and is the process by which biotin is covalently... Immunosorbent Assay) Southern/Northern blotting Cell surface labelling Attaching polypeptides to a substrate...substrate Flow cytometry/fluorescence-activated cell sorting (FACS) In situ hybridization Methods for...proteins in bacteria, yeast, insect and mammalian cells. Check out Kay et al., 2009 for methods on biotinylating...Application of avidin-biotin technology to improve cell adhesion on nanofibrous matrices.” Journal of Nanobiotechnology...

...of concept for further development. Figure 2: Mechanisms of deaminase-based adenine transversion...base editing. Molecular Cell, 84(7), 1257-1270.e6. https://doi.org/10.1016/j.molcel.2024.01.021 Tong, H.,... edit A → Y (Y = T or C) in 67.17% of screening cells (Tong, Wang, et al., 2023). Based on this promising...the base editor. Black arrows indicate natural cellular processes. Created with BioRender.com. The...Cas9 nicks the non-edited strand to encourage the cell to repair the DNA using the edited strand as a template... efficient A•T-to-C•G base editing in mammalian cells and embryos. Nature Biotechnology, 42(4), 638–650...efficient T-to-G and C-to-G editing in mammalian cells. Nature Biotechnology, 42(10), 1538–1547. https:...

...two visible blue or magenta chromoproteins (Figure 2) (Liljeruhm et al., 2018). This system is a nice confirmation... that converts sucrose into a toxic metabolite. Cells that have undergone a double crossover will not ...contain sacB, allowing them to grow on sucrose. Cells that have only gone through the single cross over...Giacalone et al., 2006). The Chen lab demonstrated excellent counter-selection which enabled very efficient...desired deletion directly into cut vector with excellent efficiency using a commercial isothermal assembly... from Harvard. Jake received his MD and a PhD in Cell and Molecular Biology from the University of Pennsylvania...

...as the protocol remains the same.    Figure 2: Microscopy-based antibody experiments should include...to detect dead or dying cells so that you can base your analysis on living cells only (for example, propidium...This tends to be most problematic when staining cells or tissues that are the same species as your primary...propidium iodide only stains dead cells). Compensation controls address the spectral overlap that occurs...Quantitative Fluorescence Microscopy.” Journal of Cell Biology, vol. 185, no. 7, Rockefeller University...

...Postdoctoral Associations around Boston - From Panel Session 2: Defining the Postdoc: An Institutional Perspective... can treat the underrepresented individuals too nicely. When the work of the underrepresented individual...

... Methods 2009; 6, 343-345. PubMed PMID: 19363495. 2. Wang JW, Wang A, Li K, Wang B, Jin S, Reiser M, Lockey...is immediately ready to transform into competent cells. The master mix of enzymes can be purchased from...mix for 1 hour, and then transform into competent cells. The success rate of this reaction is typically ...

... from damage (Abe & Lim, 2024).    Figure 2: A) When proteins (blue) interact with the air-water...epigenome editing. Cell, 184(9), 2503–2519.e17. doi: https://doi.org/10.1016/j.cell.2021.03.025.    Recombinant...tissue is identifying the cells you are studying. Antibodies can help reveal cellular identities by targeting...particles (eVLPs) offers an option to transduce cells directly with ribonucleoprotein cargo. eVLPs contain... will only be transiently present in transduced cells, limiting off-target editing. Cargo size limits ...01133-w Pandey, S., Gao, X. D., Krasnow, N. A., McElroy, A., Tao, Y. A., Duby, J. E., Steinbeck, B. J.,...site-specific integration of large genes in mammalian cells via continuously evolved recombinases and prime ...

...improvement in muscle physiological performance.  2. Trans-splicing Overview: For trans-plicing, splice...human T cells and hematopoietic stem and progenitor cells (HSPC)s to ~40% expression in the K562 cell line...gene to airway epithelial cells in mice. This vector transduced ~10% of cells, similar to the transduction...Using AAV Donor Vectors. Cell Reports 20:750–756 . https://doi.org/10.1016/j.celrep.2017.06.064  Carvalho... then independently packaged into an AAV. When a cell is co-infected with both part A and part B AAVs,... always.  The combination of serotype and tissue/cell type could have a big impact on the rate of transgene...since both of these variables influence whether cells will be co-transduced with multiple copies of both...

...signal separately for each target.   Figure 2: Examples of simultaneous multiplex IF labeling of...immunocytochemistry (ICC) when the sample is cultured cells or immunohistochemistry (IHC) with tissues), where...know if they co-localize at the same place in the cell? Or you know that the presence of several biomarkers...tissue sections, methanol- or paraformaldehyde-fixed cell samples, or other preparations. Sequential IF For...hippocampus, or counting the number of puncta in each cell. When making figures for a paper or presentation...approach for deep phenotyping and spatial analysis of cells in complex tissues. Proceedings of the National ...

...15.4%) 46 (59%) of these students were in the last 2 years of their degree. Students filled in a pre-program...professional and personal development, is essential to excelling in the modern world. Talent knows no boundaries...

...Single-chain variable fragment (ScFv) Figure 2: Comparison between the IgG antibody and scFv. ...on or in these cells: antibodies can bind to proteins on the cell surface, or the cell membrane can be...antibodies are produced by B cells. They bind to proteins on the surface of extracellular pathogens such as parasites...fusing B cells that have been extracted from animals injected with protein antigen to myeloma cells. The ... measurement of protein expression in cell lysates, whole cells, or tissue samples. Many experiments that...heterogeneous mixture - for example, cell lysate or media from cultured cells. ELISA can be used to quantitatively...same cell simultaneously as well as compare between samples. Learn more about flow cytometry. Cell and ...

...module_attribute "schema_version" is_json="true" %}{% raw %}2{% endraw %}{% end_module_attribute %}{% module_attribute...Targeted Human Stem Cell Clones Detected by Whole-Genome Sequencing. Cell Stem Cell 15:27–30 . https://...protein, in order to mitigate the ability of the cell to use an alternative ATG downstream of the annotated...process, and usually involves the need for single cell cloning and subsequent screening for successful ...standard requires not one but two rounds of single cell cloning – as a control, one should revert the edit...passenger variant that came along with the single cell clone (although this is rarely done). When targeting...levels of modification. Whole-genome sequencing of cells modified by CRISPR indicates that the consequences...

...effective recovery of knock-ins, but did result in 1-2 kilobases of additional sequence being introduced....experiments possible in a wide range of organisms and cell lines. In C. elegans, one of the model organisms... efficiency. Finally, as shown in Drosophila S2 cells (Böttcher et al., 2014), inactivation of non-homologous... community. Developments in mammalian and yeast cells using modified Cas9 proteins to regulate gene expression...homologous recombination donors in cultured Drosophila cells. Nucleic Acids Research 42:e89–e89 . https://doi.org...

...experience in cancer research After graduating with a 2:1 BSc in Molecular Biology (roughly a B average in...that, when those contracts fall through or are cancelled, then redundancies can be made within the company...

...ask new and interesting questions in your own lab. 2. Seek multiple mentors during your postdoc At the ... optimization of fluorescent proteins,  and live cell imaging approaches. Erik is also an avid long distance...

...biology 14.6 (2004): 757-764. PubMed PMID: 15582400. 2. Tomkinson, Alan E., et al. "DNA ligases: structure...ligase bound to nicked DNA-adenylate." Molecular cell26.2 (2007): 257-271. PubMed PMID: 17466627. 25. Luo...electrophoresis as a sensitive, high-throughput method to accelerate characterization of nucleic acid metabolic enzymes...

... overnight culture to 500 μL of 50% glycerol in a 2 mL screw top tube or cryovial and gently mix. Notes...bacteria, preventing damage to the cell membranes and keeping the cells alive. A glycerol stock of bacteria... and you will not need to obtain more competent cells and retransform. Bacteria on an LB agar plate can...

...10μL reaction for 5X buffer) 0.5-1μL T4 DNA Ligase H 2 O to a total of 10μL Notes: If the DNA concentrations...complete plasmid can be transformed into bacterial cells for propagation. The majority of ligation reactions...Interpretation Uncut vector - Checks viability of competent cells and verifies the antibiotic resistance of the plasmid...

...nonspecific cation influx into the cell when stimulated. Channelrhodopsin-2 (ChR2), the first widely adopted... (Phy) interaction 660 Cryptochromes Cryptochrome 2 and CIB1 Blue-light–mediated induction of protein ...interactions based on Arabidopsis thaliana cryptochrome 2 and CIB1 450 LARIAT Inhibits protein function by reversibly...customizable TALE DNA-binding domain with cryptochrome 2 protein and its interacting partner CIB1 to control...transcription effectors OPTOSTIM PHR domain2 of cryptochrome 2 (Cry2) from Arabidopsis thaliana is fused to truncated...control of biological systems. Nat Rev Methods Primers, 2 , 55. https://doi.org/10.1038/s43586-022-00136-4 PMID... with single-spike temporal resolution. PloS One, 2 (3), e299. https://doi.org/10.1371/journal.pone.0000299...

...use of pAdEasy-2 (Addgene #16401) can increase the capacity of the rAdV vector. pAdEasy-2 does not contain...gene and cell-based therapies in the new era of personalized medicine . Genes & Diseases, 4 (2), 43–63.... incorporate additional genomic deletions. Figure 2: First-generation rAdV vectors. Created with BioRender.com...and have to be handled at biosafety level two (BSL-2). They are highly immunogenic and can trigger a strong...adenoviruses using the AdEasy system . Nature Protocols, 2 (5), 1236–1247. https://doi.org/10.1038/nprot.2007.135...Adenoviral vector vaccine platforms in the SARS-CoV-2 pandemic . NPJ Vaccines, 6 (1), 97. https://doi.org...backbone plasmid that lacks E1 and E3. pAdEasy-2 Adenoviral backbone plasmid that lacks E1, E3, and E4. Useful...

...DNA concentration alone. One method is to conduct 2 ligations for each plasmid you are trying to create...use electro-competent cells instead of the more common chemically-competent cells. The number of bacterial...might need to express YGOI in cultured mammalian cells. The problem is that the only version of full-length... manufacturer’s instructions for your competent cells. For most standard cloning, you can transform 1-...1-2μl of your ligation reaction into competent cells such as DH5alpha or TOP10. If using much less total...colonies, you might want to use higher competency cells. Additionally, if your final product is going to...

...assembled into a Level 2 vector, forming a functional genetic circuit. Level 2 vectors are often designed...of three sets of cloning vectors (Level 0, 1, and 2) which can be used in successive assembly steps. Before... iterations of assembly. Combining multiple Level 2 vectors permits the creation of even more complex ... 16 RBS strength variants, 8 tag-compatible RBSs, 2 secretion tags, 6 CDSs, 9 terminators, 4 nonfunctional...of multigene constructs. PLoS One . 2011 Feb 18;6(2):e16765. doi: 10.1371/journal.pone.0016765. PMID: ...assembly of cellular circuits using Golden Gate-based cloning methods for use in mammalian cells. These can...MoClo Yeast Toolkit (YTK) to optimize secretion and cell surface display of proteins of interest. Yeast Secrete...

...muscarinic receptor and has not been used in vivo. Figure 2: DREADDs, their ligands, and signaling properties.... Reference Rq(R165L) Human M3 muscarinic Arrestin-2/-3 CNO* Increase Arrestin translocation Arrestin signalling...hM1Dq hM5Dq Human M3 muscarinic G αq CNO* Increase Ca 2+ Neuronal burst firing Armbruster et al., 2007 rM3D...channels, their effect, and outcome in neurons. Table 2: Descriptions of PSAMs and their activity in neurons...FRET-modified bioluminescence. Neurophotonics 11 (2), 021005. https://doi.org/10.1117/1.NPh.11.2.021005...improved coupling efficiencies. Neurophotonics, 11 (2), 024208. https://doi.org/10.1117/1.NPh.11.2.024208...targeted to specific tissues, cell types or even subcellular regions of a neuron. Cell-specific expression of...

...intact. First-generation plasmids include (Figure 2): Transfer plasmid — contains transgene and wild-type...envelope removed Envelope plasmid — contains env Figure 2: First-generation lentiviral plasmids. Created with...plasmid 1 — contains gag and pol Packaging plasmid 2 — contains rev Envelope plasmid — contains env Third-generation...The inside out of lentiviral vectors. Viruses , 3 (2), 132–159. https://doi.org/10.3390/v3020132 PMID: ...Infectivity and Integration Targeting. PLoS ONE , 2 (12), e1340. https://doi.org/10.1371/journal.pone.0001340...features of lentiviral vectors. Human Gene Therapy , 24 (2), 132–142. https://doi.org/10.1089/hum.2012.229 PMID...production cell line, typically human embryonic kidney 293T (HEK293T) cells. Standard HEK293T cells are sufficient...

...DNA concentration alone. One method is to conduct 2 ligations for each plasmid you are trying to create...use electro-competent cells instead of the more common chemically-competent cells. The number of bacterial... manufacturer’s instructions for your competent cells. For most standard cloning, you can transform 1-...1-2μl of your ligation reaction into competent cells such as DH5alpha or TOP10. If using much less total...colonies, you might want to use higher competency cells. Additionally, if your final product is going to...

...Haploid Embryonic Stem Cells Carrying a Guide RNA Library. Cell Stem Cell . 17 (2), 221–232. PMID: 26165924...mutant cell must be validated experimentally. Browse Plasmids: Double-Strand Break (Cut) Figure 2: Overview...used to transduce your experimental cell population (Figure 8E). In a 2-vector system, you’ll need to either...of CRISPR-Cas9 with Bacteriophage Proteins. Cell , 168 (1–2), 150-158.e10. PMID: 28041849 Sakuma, T., Nishikawa... regulation of transcription in eukaryotes. Cell . 154 (2), 442–451. PMID: 23849981 Gilbert, L. A., Horlbeck...Transcriptional Programs with CRISPR RNA Scaffolds. Cell . 160 (1–2), 339–350. PMID: 25533786 Cas9 alternatives...single RNA-Guided endonuclease of a Class 2 CRISPR-CAS system. Cell , 163 (3), 759–771. PMID: 26422227 Zetsche...

...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...

...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...

...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...

...types: positive screens and negative screens (Figure 2). Both types of screen employ a selection method relevant...planning to conduct a pooled library screen. Figure 2: A simplified outline for using lentiviral gRNA libraries...200x the number of cells as plasmids in the TKO library. Using large numbers of cells minimizes the effects...selection Most cells die or do not pass selection (in the case of a reporter) Sort “winning” cells Sequence ...library Apply selection; most cells will survive Perform NGS on the surviving cells Compare NGS results between...identify those cells that do not survive the selection mechanism. You’ll infect two sets of cells and subject...libraries, plasmid DNA can be delivered directly to the cells of interest. With others, notably pooled lentiviral...

...positions from the transcription start site (TSS) (Figure 2). The Pribnow box (TATAAT) is located at the -10 position...recognizes different core promoter sequences. Figure 2. Basic structure of a bacterial promoter. TSS = transcription...element controls the rate of transcription. Bacterial cells contain sigma factors that assist the RNA polymerase...other promoters can drive expression in specific cells or under different conditions. Promoter Expression...ribosomal RNA (rRNA), which is a main component of a cell’s ribosome structure. Ribosomes are the site of protein...

...specific control of activity. Read More CRISPR Class 2 C lustered R egularly I nterspaced S hort P alindromic...receptors to selectively control and investigate cell signaling pathways. Receptors are remotely controlled...and genetic engineering to measure and manipulate cells (frequently neurons) and their governing biomolecular...detect, measure, and control molecular signals and cells in order to understand their functions. Read More...

...range of infectivity for different cell types (known as tropism). Figure 2: Gamma-retroviral plasmids. Created...gamma-retroviral vectors, you need three plasmids (Figure 2): Transfer plasmid — contains transgene, sgRNA, or...-activated cell sorting (FACS) is used to sort cells expressing GFP and later, sorted cells are expanded...actively dividing cell types, whereas gamma-retroviruses can only infect mitotically active cell types. This...Using Packaging Cell Lines Gamma-retroviral vectors can be packaged using helper-free cell lines (Figure...-retroviral packaging cell line developed by Garry Nolan at Stanford. These cell lines contain gag-pol...and rat cells), or an amphotropic envelope, Phoenix-AMPHO (for the infection of mammalian cells). Using...

...unique restriction enzyme recognition site (Figure 2). These elements allow for the propagation of the ...elements are summarized in the table below. Figure 2: General structure of a plasmid. Plasmid Element Description... plasmid DNA is replicated upon cell division, and each daughter cell receives at least one copy of the...segregated to the resulting daughter cells. These daughter cells are called clones, since they contain...non-coding RNA (ncRNA) and play important roles in cellular processes and gene regulation. Regions that code...techniques enable the isolation and purification of cellular components, such as DNA, but in practice this ..., which take up plasmids and amplify them during cell division. Because bacteria are easy to grow, divide...

...Read more in our Gateway Cloning blog post . Figure 2: Summary of Gateway cloning. Created with BioRender.com...homology. While this approach doesn't work in other cell types, it can be a major time- and cost-saver for...