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...Plasmids need antibiotic resistance genes so that you can specifically isolate bacteria containing your...action of the most common antibiotic resistance genes and which applications are best (or worst) for each...plasmid of interest, preventing this issue. The genes and their antibiotics   Ampicillin Ampicillin, often...antibiotic due to stable integration of resistance genes in their genomes. If you need a plasmid that can... Zeocin-induced DNA cleavage in human cells. Mutagenesis, 20(2), 111–114. https://doi.org/10.1093/mutage...

...breast and ovarian lines commonly used in lab. It is recommended you check your cell lines for HR-related ...covered in this blog, with tips and tricks for harnessing the homology-directed repair pathway, designing...surprise then that these edits take more planning and finesse then their knock-out counterparts. To kick off ...experiment today! Get ready to identify knock-in clones by reading up on validating genome edits to find...

...biological disciplines and scientists employ GFP for a wide number of functions, including: tagging genes for...Bioluminescence and fluorescence from proteins such as Green Fluorescent Protein (GFP) has likely existed...began engineering new versions of GFP through mutagenesis in order to improve its physical and biochemical...Click here to view Addgene’s collection of empty backbones for constructing fluorescent fusions. Transcription...

... help you verify the ends of the insert. Longer genes may require internal primers that anneal to a specific...

...digests needed in molecular biology, and thaw cell lines. There are many different types of water baths that...

...To deliver genes using lentiviral vectors, you need an envelope protein on the virus’s surface and a ...stomatitis virus, HIV-2, and human foamy virus in cell lines expressing galactosyl(alpha1-3)galactosyl (alphaGal... were less stable than viruses produced in cell lines that do not express these sugars. (Takeuchi et al...study the spike protein’s role in infecting cell lines without using intact SARS-CoV-2 virus. Scientists...find tables and information on different viral membranes, tropism, and advantages and disadvantages: Table...

...AAV enhancer plasmids with high specificity and brightness, now available at Addgene!   {% module_block ... those specific cell types to express exogenous genes — a feature the AIBS has capitalized on. ...containing enhancers with the highest specificity and brightness. You will find enhancers to target many different.... A suite of enhancer AAVs and transgenic mouse lines for genetic access to cortical cell types. Cell...., 288–295. https://doi.org/10.1038/nrg3458 Ray-Jones, H., & Spivakov, M. (2021). Transcriptional enhancers...

... propagation of themselves and their associated genes. These systems better ensure that plasmids are passed...death. The team used this cassette to knock out genes, knock in a GFP cassette, and introduced site-directed... (hok), a long-living toxin that damages cell membranes; Suppression of killing (sok), a short-lived RNA...proof of concept, they tested a lux-mediated bioluminescence reporter in vivo in the absence of selective... Ogura, Teru, and Sota Hiraga. "Mini-F plasmid genes that couple host cell division to plasmid proliferation...

... similar brightness - even though in the most popular pair, CFP has five-fold less brightness than YFP...Master. Note: BRET= Bioluminescence Resonance Energy Transfer CRET= Chemiluminescence Resonance Energy Transfer...that can affect FRET measurements include: the brightness of a fluorophore pair, donor:acceptor stoichiometry...

...Isolation and Concentration (MagIC)-cryo-EM that combines enrichment and imaging of proteins captured on...with a GFP nanobody (Figure 1). This enrichment streamlines the purification and minimizes sample loss while...loss. Ci Ji Lim’s lab found that adding Late Embryogenesis Abundant (LEA) proteins, small proteins that... to partially correct mouse models of genetic blindness (An et al., 2024) — the first demonstrated use...). Efficient site-specific integration of large genes in mammalian cells via continuously evolved recombinases...

... are essential for processes such as gluconeogenesis, lipogenesis, amino acid metabolism and energy transduction...reactive and forms a link with nearby nucleic acids (McInnes et al., 1990). Photobiotinylation is sometimes ...18988027. PubMed Central PMCID: PMC3223083. 6. McInnes, J. L., Forster, A. C., Skingle, D. C., & Symons...Biotinylation ThermoFisher Scientific: Biotinylation Guidelines  ...

...Repair is Used to Edit Genes Read How Non-Homologous End Joining is Used to Delete Genes Additional Resources...exons), but much of the sequence appears to be nonessential. Within the mutational hotspot for dystrophin... cells expressed functional dystrophin. Editing genes in vivo is of course much more difficult than in...these different mutations. For each therapy, effectiveness and off-target risk must be evaluated separately...

...plasmid yield and/or DNA quality. Table 1 below outlines a few of the more common genetic changes found...encodes proteins needed for bacterial conjugation. Genes listed on F´ are wild-type unless indicated otherwise...lac Lac operon mutations Blue/white screening of clones   lacIq lac repressor overproduced, expression...transformation efficiency deoR constitutive expression of genes for deoxyribose synthesis Allows uptake of large...)]  Note: Inactivating mutations in specific genes are signified with a minus sign (-) as is typically...

... and may provide counter-selection for negative clones as in the example shown here. The manufacturer ...

...Finally, as Pamela Hines of AAAS points out on the panel, irreproducibility undermines public trust in science...Last Wednesday we worked with the Harvard GSAS Science Policy Group to organize a Minisymposium on Reproducibility...) - Panel Alex Tucker (Ginkgo Bioworks) Pamela Hines (Senior Editor at Science) Edward J. Hall (Professor...journalists and scientific experts discussed by Pamela Hines Minisymposium on Reproducibility Booklet - Including...

...involves making a large sgRNA library to thousands of genes and targeting and sequencing millions of cells. ...cell line, with the goal of isolating single cell clones to use for multiple downstream experiments. You...certain Cas enzymes are less compatible with the bulkiness that accompanies mispairing of bases. To reduce... characterization of, isolating 2 or 3 distinct clones to perform key experiments on is a good way to ... distinct off-target sites, eliminating its effectiveness as a control. Since off-targets typically confound...

...al., 2009), and at high copy numbers (Williams, Carnes, and Hodgson, 2009). More research is ongoing to...1566524020666200427110452. PMID: 32338220. Crozat E, Fournes F, Cornet F, Hallet B, Rousseau P. Resolution of...Levene SD. Analysis of DNA topoisomers, knots, and catenanes by agarose gel electrophoresis. Methods Mol Biol...0092-8674(84)90060-6. PMID: 6323019. Williams JA, Carnes AE, Hodgson CP. Plasmid DNA vaccine vector design... JA, Luke J, Langtry S, Anderson S, Hodgson CP, Carnes AE. Generic plasmid DNA production platform incorporating...

...many of which are used to deliver Cas9 and other genes to mammalian cells. For this reason, the top requested...means researchers who use VSV-G can deliver their genes of interest to a variety of cells and study their...Skimming the list, one gets a sense of the broad usefulness of pMD2.G and VSV-G. This plasmid has been used... of the secretome, ubiquitin ligase, cellular senescence, autophagy, CRISPR, and so much more (find the...

...edits and to perform functional screens to identify genes involved in a phenotype of interest. This blog post...protein-coding capacity of a locus. When using S. pyogenes Cas9 (SpCas9), potential target sites are both...Veres et al., 2014). When working with single-cell clones, the authors note that “clonal heterogeneity may...obstacle to the generation of truly isogenic cell lines than nuclease-mediated off-target effects.” Further... CRISPR-Cas9 and TALEN Targeted Human Stem Cell Clones Detected by Whole-Genome Sequencing. Cell Stem ...

...societal implications even before the discovery of genes themselves.  How has scientific progress in genetic...and vice versa? We draw a parallel between the timelines of the scientific advancements and the noteworthy...to stars in space ships and the rise of killer machines that exterminate entire humanity were the two ...that results in gene knockouts.   Star Trek: Into Darkness (2013) which is the remake of the space-melodrama...Hugo De Vries: from the theory of intracellular pangenesis to the rediscovery of Mendel. Comptes Rendus ...