We narrowed to 672 results for: cell

...development. Single cell sequencing-based lineage tracing Recently, gene expression in all cells of the developing...proportions of otherwise normal cell types are produced, rather than novel, abnormal cell types. Sequencing-based...applications. For example, specific tissues such as blood cells or neurons can be fluorescently labeled, allowing...divisions, and movements of each of the thousands of cells in the developing zebrafish embryo have been tracked...developing zebrafish embryo has been mapped using single cell RNA sequencing approaches (Harland, 2018). Several...sequenced zebrafish genome. One study profiled 38,731 cells from 694 embryos over multiple stages and reconstructed...reconstructed the patterns of gene activity and cell divisions that coordinate development of the body plan...

...relied on for transducing cells: galactose and the AAV receptor (AAVR). Cell lines lacking galactose were...Concise review: stem cell antigen‐1: expression, function, and enigma." Stem cells 25.6 (2007): 1339-1347... Wilson labs determined LY6A, also known as stem cell antigen-1 (SCA-1), is the receptor used by PHP.B...commonly used to enrich adult mouse hematopoietic stem cells. Its ligand(s) has not been identified. While LY6A...out of Ly6a in primary C57BL/6J brain endothelial cells decreased transduction rates while overexpression...overexpression of the C57BL/6J version of Ly6a in 293T cells enhanced PHP.B transduction rates. The group also determined...PHP.B, virus still reached the brain endothelial cells of these mice, although at lower levels than in ...

...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...two weeks to allow differentiation to occur. CD8+ cells were then isolated and their genomic DNA sequenced...also be applied to the differentiation of other CNS cell types, including microglia, astrocytes, and oligodendrocytes...

...no detectable effect on cell viability – meaning there was no measurable cell toxicity – using Cas7-11...such as mouse embryonic stem cells and glioblastoma cells. PASTE Development and discovery The first ...observed in non-dividing cells, meaning that this system wasn’t dependent on the cell’s intrinsic machinery... shRNA and Cas13 systems in a range of mammalian cell lines. However, in those same comparisons, the AbuGoot...11. This included tests done in cell lines known to be more sensitive to toxicity from RNA editing, such...integrases with efficient integration activity in human cells. Moreover, they optimized the specific attachment... of plasmids, works in non-dividing and primary cells, and can be easily programmed for new genes. In ...

...synthesized inside of what is called the “target cell” (or host cell) by the viral protein reverse transcriptase...reverse-transcribed DNA into the host cell’s genome and then use cell machinery for transcription and translation...1971). Noninfectious ARK mouse embryo cell lines in which each cell has the capacity to be activated to ..., interested in transferring genes to mammalian cells (i.e. transferring DNA), yet retroviruses are RNA-containing...separate set of instructions in the same producer cells as the virus. Because it is made at the same time...viral genes. It is derived from the host/producer cell membrane, and acquired as an envelope during exocytotic...particles do once they come in contact with target cells (Fig. 3).     Figure 3: The life cycle of...

...spread. They can be inherited by daughter cells during cellular reproduction, similar to the host’s chromosome...elements in the human gut. Cell, 187(5), 1206-1222.e16. https://doi.org/10.1016/j.cell.2024.01.039 Garcillán-Barcia...bacteriophages), or conjugation (direct transfer from one cell to another). Bacteria need special fertility genes... of a plasmid benefiting its host. For example, cells with a penicillin resistance plasmid can grow in...also carry the necessary resistance genes to help cells survive their own secretions. Pro tip! Historically...bacteria invade hosts, including attaching to host cells and evading the host immune system, and those that...used routinely in biotechnology to transform plant cells. Degradative plasmids: microbial hardiness There...

...This novel tool is practical in cell and organ culture, as well as cell lineage studies. Eisenhoffer and...CreLite for cell tracing using the transgenic zebrafish model, ubi:zebrabow. In this system, cells express...fast-splicing split intein and co-infected into the cell. Intein splitting removes any exogenous sequences...split into two plasmids and co-transfected into cells for reconstitution. Image from Levy et al., 2020...red-light inducible optogenetic tool for precision in cell lineage tracing Article contributed by Erin Sanders...fluorescent proteins controlled by Cre recombination. Each cell can have multiple copies of the transgene that recombine..., such as the brain, eye, muscle and epithelial cells, as well as in different stages of development, ...

...plasmid. Envelope proteins determine which types of cells a virus can infect. The VSV-G envelope protein enables... enables this virus to infect many different cell types so it is useful in a wide range of applications... The 2A-GFP allows for screening of transfected cells. Find pSpCas9(BB)-2A-GFP (PX458). lentiCRISPR v2...deliver Cas9 and your gRNA of interest into mammalian cells. This vector was the top requested plasmid of 2015... popular in genome engineering. For use in human cell lines, the Zhang lab codon-optimized SpCas9. Using...puromycin cassette that makes it less efficient in some cell lines. pSpCas9(BB)-2A-Puro (PX459) V2.0 was made...capable of inducible expression of shRNA in target cells by the addition of tetracycline/doxycycline to the...

...dodge cellular variation observed in mammalian cells is to tag an endogenous gene. By imaging cells (or ...mammalian cells within the dish. This is caused by the stochastic nature of plasmid uptake by cells. To correct...intended application. The three key properties for live cell imaging that will be treated in this series of posts...Determining practical brightness When a dish of mammalian cells is transiently transfected with a plasmid containing...allows for a precise determination of brightness in cells, under realistic experimental conditions. By determining...al (2017) and Heppert et al (2016). In mammalian cells this would be possible with CRISPR/Cas-based genome...efficiency of the fluorescent protein in the host cell, which is a key determinant of the practical brightness...

...expressed in cells that express cre recombinase. Typically, this feature is used to attain cell-specific ...CRISPR applications and can be expressed from your cell line of choice after transduction with this virus...blast virus can be used to generate Cas9-expressing cell lines, which can then be used both for screens (...viral prep to create a single stable Cas9-expressing cell line and then transduce this line with lentiviruses...using lentiCas9-blast to generate Cas9-expressing cells. To ensure that this growth rate effect doesn’t ...activating particular signaling pathways in specific cell types (typically neurons). While all of these viruses...transduction and transgene expression across different cell types (Aschauer et al., 2013). Expression of hM4D...

...imaging with single-cell resolution using chemical cocktails and computational analysis." Cell 157.3 (2014):... of the XFP genes is introduced at ~2 copies per cell per color, following a Poisson distribution. To ... optimal expression of XFPs. Any longer and the cells may start to show morphological abnormalities due...XFP genes have to be introduced at ~2 copies per cell per color to achieve the highest color variations... wait too long, as too much XFP is toxic to the cells. Inadequate expression levels of XFPs - Reduce the...video below depicting dendrite wiring of mitral cells.   Many thanks to our guest bloggers Richi Sakaguchi...circuitry with an index-optimized clearing agent." Cell reports14.11 (2016): 2718-2732. PubMed PMID: 26972009...

... disease, cell type or drug treatment can browse through screens performed in relevant cell lines or conditions... currently houses over 890 CRISPR screens in 629 cell lines, each manually curated and annotated from ...feasibility and power of CRISPR-based library screens in cell lines from humans and other species. By accurately... screens performed in 26 different breast cancer cell lines. Screenshot updated 2024. Each screen ...screen is captured by BioGRID curators, including the cell line/tissue type, enzyme system, sgRNA library and...example phenotype screens with read-outs other than cell viability. The following video provides an in-depth...

...Localization within cells Fluorescence microscopy Visualize components of individual cells Separation/identification...fluorescently-labeled components Fluorescence-activated cell sorting (for cells); fluorescent HPLC (proteins or other molecules...biosensor Before expressing a biosensor in eukaryotic cells, think about the best place for the biosensor to...average amount of fluorescence across a population of cells (i.e. in a plate well). Use to track a process (... molecules) Isolate or quantify cells, proteins or other molecules that have been labelled with your fluorescent...be expressed. Are you looking at an intracellular or extracellular molecule? By fusing a biosensor to ...by Olivier Pertz’s lab. Other biosensors track cellular characteristics like pH, redox state, and voltage...

...recombination varies for different constructs and cell types. Second, Cre may induce recombination at pseudo...standard UAS/GAL4 system is toxic to proliferating cells. A Cre-estrogen receptor ligand binding domain-fusion...recombination efficiency. When expressed in neuronal cells, each line showed broad recombinase-dependent RFP...these recombinases were also active in mammalian cells. In experiments, B3 and KD were shown to be functional...functional in cultured Chinese hamster ovary (CHO) cells, as well as in cortical neurons in layer 2/3 of ... damage induced by Cre recombinase in mammalian cells. Loonstra A, Voojis M, Beverloo HB, Al Allak B, ...recombinase toxicity in proliferating Drosophila cells by estrogen-dependent activity regulation. Heidmann...

...eight millimeters – not single cells (yet) but small clusters of cells. Looking ahead What’s next, you...fluorescent proteins to observe what’s happening inside cells for at least a couple of decades. Green is the classic...and tracking of proteins, organelles, and living cells in a spatiotemporal manner, which is not possible... vivo. “One of the reasons we can’t localize one cell at a depth of one centimeter with fluorescence is...on the other hand, are larger than the size of a cell, so they scatter much less. As a result, ultrasound...brighter, more photostable, smaller, and less toxic to cells. They’d like to make near-infrared fluorescent proteins...

...lipopolysaccharide of the cell wall of Gram negative bacteria like E. coli that affects tissue culture cell line viability...introduces transformation grade plasmids into a bacterial cell. Transformation grade DNA  Transformation is ...transformation of chemically and electrocompetent cells is minimal (picogram range). It can be extracted...Figure 3: Need to introduce plasmids into mammalian cells? Use transfection grade DNA. Transfection is ...process of inserting an external plasmid in mammalian cells and it will likely require micrograms of plasmid...

... them into your cells, you can simply look for blue colonies to determine which cells have taken up the... mScarlet as the donor in HeLa cells.   Find ShadowR for mammalian cell expression here   Now that you... that the cellular fitness cost varied greatly between chromoproteins. In some cases, cells containing...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...resonance energy transfer (FLIM-FRET) in living cells (Figure 4). They created ShadowR by modifying the...

...ancestry and germline sequence variation in cancer cell lines took them down unexpected paths. Though this...researchers do large-scale CRISPR screens in cancer cell lines, looking to uncover genetic dependencies of...to see if they could identify the ancestry of the cell lines (European, East Asian, Indigenous American...or South Asian) used in the screens based on the cells’ germline genetic variations. They then wanted to...with each ancestry group. Since about 90% of the cell lines used in DepMap CRISPR screens are of either...groups for 994 of the Cancer Dependency Map cancer cell lines (left). Heatmap indicating the breakdown of...CRISPR guides, short pieces of RNA that bind to a cell’s DNA, telling the Cas machinery where to cut.  ...

...Depletion in Human Cells by Auxin-Inducible Degron Tagging with Short Homology Donors.” Cell Rep.15(1) (2016...and function of cellular systems. FPs have been used to track protein localization, cell structure, intracellular...to fluorescently tag markers of cellular structures in mammalian cells. These plasmids use fluorescent...loci in other model systems, including mammalian cells. CRISPaint (CRISPR-assisted insertion tagging)...C-terminal tag fusions of endogenous genes in human cells. To use CRISPaint, users need 3 vectors:  1) a gRNA...also be introduced directly into a Drosophila S2 cell line stably expressing Cas9. This PCR toolkit offers...selection. Researchers at the Allen Institute for Cell Science recently developed a collection of plasmids...

... into AAV particles for delivery into mammalian cells, especially when fused to effector proteins for ..., base editing, and genome editing in mammalian cells. The “V4” version (named CasMINI) had the highest...an active nuclease.  Read the paper in Molecular Cell Find the CasMINI plasmids Efficient CRISPR editing...effective in genome editing in both bacteria and human cells using various delivery methods, including plasmid...mRNA or Cas9 mRNA and sgRNA, in human and mouse cells. They further develop a version of SEND for gene...modular system for delivering gene editing tools into cells. Image adapted from Segel et al. 2021.   Prime...correct mutations in a variety of target genes and cell types. Their Nature Biotechnology paper showcases...