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We narrowed to 130 results for: yeast expression

Showing: 41 - 60 of 130 results

  1. Protocol - pLKO.1 – TRC Cloning Vector

    Type
    Protocol
    ...replication-incompetent lentiviral vector chosen by the TRC for expression of shRNAs. pLKO.1 can be introduced into cells...Human phosphoglycerate kinase promoter drives expression of puromycin. Puro R Puromycin resistance gene...pLKO.1 vector, producing a final plasmid that expresses the shRNA of interest. C.1 Recommended Materials...HIV-derivative viruses. For transient knockdown of protein expression, you may transfect plasmid DNA directly into ...into the target cells. The shRNA will be expressed, but the DNA is unlikely to be integrated into the host...will allow you to select for cells that stably express your shRNA of interest. E.1 Recommended Materials...allow you to select for cells that are stably expressing your shRNA of interest. F.1. Recommended Materials...

  2. What the HEK?

    Type
    Blog Post
    ...derivatives of HEK-293, the most commonly used line expresses SV40 large T antigen and is thus dubbed HEK-293T...packaging cells’ for viral production because they express the SV40 large T-antigen. This allows them to replicate... with other protein production systems, such as yeast. HEKs can also grow in suspension, which allows ...receptors and ion channels are often exogenously expressed and studied in HEKs due to the cell line’s ability...produce these proteins and its relatively low expression of native channels.  No matter what your research...factors that could skew research results (gene expression, growth rates, etc.). Thus, it’s important to...

  3. Plasmids 101: What is a plasmid?

    Type
    Blog Post
    ...Vital component for expression vectors: determines which cell types the gene is expressed in and amount of...exist naturally in archaea and eukaryotes such as yeast and plants. In nature, plasmids provide one or more...sites allowing for the easy insertion of DNA. In expression plasmids, the MCS is often downstream from a ...Generally, scientists use plasmids to manipulate gene expression in target cells. Characteristics such as flexibility...common plasmid types include cloning plasmids, expression plasmids, gene knock-down plasmids, reporter ...

  4. Quick Guide to Working with Drosophila Part 2: Controlling Gene Expression in Flies with Gal4/UAS

    Type
    Blog Post
    ...gene expression. Thus, genes and siRNA encoded downstream of the UAS sequence are only expressed when ...when Gal4 is expressed. Gal4 expression can be regulated by any chosen promoter, and Bloomington’s Stock ...fly lines expressing Gal4 under a variety of promoters resulting in tissue specific expression. To express...genes and wanted to express them within a specific neuronal compartment, you could express them individually...modular expression systems exist (eg. LexA/LexAop and Cre/Lox systems) in case you want to express one gene...system to knock down gene expression, rescue knock out mutations, and overexpress viral genes. Additionally...with Drosophila, you’ll learn how to maniupate expression of your favorite gene (YFG) in flies. Read the...

  5. Fluorescent Proteins 101: Photoactivatable Fluorescent Proteins

    Type
    Blog Post
    ...protein‐tracking studies in the budding yeast Saccharomyces cerevisiae." Yeast 25.9 (2008): 651-659. PubMed PMID...investigate protein localization, interaction, and expression. However, one caveat of FP-protein fusions (FP-chimeras...determine specific protein turnover or temporal expression using standard FP-chimeric proteins. Photoactivatable...applications PA-FPs have been developed for use in yeast, as parts of biosenors, and much more. Do you have...

  6. Plasmids 101: Mammalian Vectors

    Type
    Blog Post
    ...components expressed in trans within the cell for effective replication. Cell lines expressing the Epstein–Barr... their genomes. Unlike plasmids used for expression in yeast and bacteria, plasmids used for stable transfections...straightforward and the resultant cells can either express the plasmid DNA transiently (similar to bacteria... easily monitor transfection efficiencies and expression levels in the cells. Ideally, the chosen reporter...reporter is unique to the cell, is expressed from the plasmid, and can be assayed conveniently. A direct... sufficient for the transfected plasmid to be expressed transiently. Since the DNA introduced in the transfection...to the host genome, the cells will continually express the genetic traits of the foreign material and ...

  7. Plasmids 101: Inducible Promoters

    Type
    Blog Post
    ...gene, it therefore determines the timing of gene expression and largely defines the amount of recombinant...lac inducible system is found in many bacterial expression vectors. Negative inducible promoter pBad is ...Temperature inducible promoters Temperature sensitive expression systems are typically less leaky than chemically...chemically induced promoters; they show near-zero expression at regular temperatures but can be induced by ... promoters, in which a gene of choice is only expressed following exposure to a brief heat shock. In the...promoters Light is another way to activate gene expression, and two-component systems used in synthetic ...transcription from phage promoter pR, preventing expression of a reporter gene. When light is present, YFI...

  8. Plasmids 101: Blue-white Screening

    Type
    Blog Post
    ...operon repressor, thereby allowing lac expression). When expressed, the β-galactosidase enzyme can break... that encodes for the enzyme β-galactosidase. Expression of the lac operon is induced by lactose, and ...enough time for any intact β-galactosidase to be expressed and process x-gal into blue pigment (16-20 hours...Positive selection vectors encode a gene which, when expressed, is lethal to the cell. Cloning fragments are ...Post on Common Lab E. coli Strains Learn to Use Yeast Two Hybrid Systems Browse All of Our Plasmids 101...You Browse Addgene's curated list of Bacterial Expression Systems Find Empty Backbones for Your Experiments...

  9. Hot Plasmids - September 2019 - CRISPR, Auxin Inducible Degradation, MerMAID Channelrhodopsins, and Genome Looping

    Type
    Blog Post
    ...tagging proteins with a 7kD degron called mAID and expressing the essential plant protein TIR1 in non-plant..., the Kanemaki lab was able to translate AID to yeast, as well as mammalian and human cell lines (Nishimura... require long time scales with constitutively expressed proteins that are not inducible. To further develop...observe loop formation as well as increased Zfp462 expression within 4 hours of blue light induction (6-fold...

  10. Hot Plasmids - June 2020 - Barcoded CRISPR Library, Sparse Cell Labeling, Calcineurin Reporter, and DNA Staining Dye Alternative

    Type
    Blog Post
    ...the stop cassette is removed, the effector is expressed. By truncating one of the recombination sites ...destabilized GFP reporter for calcineurin activity in yeast Article contributed by Leah Schwiesow Calcineurin...calcineurin-dependent response elements (CDREs) and drives gene expression. Because calcineurin is required for virulence... single molecules in real-time. Find plasmids expressing tTALE-FP Shin et al., Sci. Rep. 2019. https:/...

  11. Hot Plasmids - March 2020 - base editors, GEVI, MoClo, and optogenetics

    Type
    Blog Post
    ...determined protein expression levels from different promoters, and inhibited gene expression using CRISPRi....transduced at levels similar to that of a single AAV expressing GFP. The lab tested the base editors in vivo ... for use in model organisms such as E. coli and yeast. Because this approach uses standardized parts, ...increases temporal and spatial precision of gene expression and reduces toxicity as compared to previous ...zebrafish model, ubi:zebrabow. In this system, cells express various fluorescent proteins controlled by Cre ...depending on the combination of fluorescent proteins expressed. CreLite-mediated recombination was found in both...

  12. Plasmid Tools for Microbiome Studies

    Type
    Blog Post
    ...can also detect expression of toxin mRNA from Clostridium difficile. As mRNA expression cannot be identified...Gate Assembly. They built this toolkit upon the Yeast Toolkit by replacing the backbone vector’s high-...resistance markers, promoters, and inducible gene expression systems in bee gut microbiome species such as... or a bacterial microcompartment protein. The expression of this fusion is under control of the arabinose-inducible...Cells are first induced with arabinose to begin expressing the SAP-RFP fusion. Then, the cells are washed...

  13. Plasmids 101: Terminators and PolyA signals

    Type
    Blog Post
    ...Plasmids designed to express genes in a given host cell type are generally broken down into two broad...on promoter strength as a determinant of gene expression levels, the terminator also plays an important...variability in RNA half-life, and ultimately gene-expression. Polyadenylation is the post-transcriptional ...terminators are not usually employed in plasmid-based expression systems, so these will not be detailed here, ...provided at the end.  Nearly all common bacterial expression plasmids use Rho-independent terminators, which...common terminator will suffice. Many commercial expression vectors use double terminators to reduce unwanted...couple termination with polyadenlyation. Mammalian expression plasmids are primarily used to create mRNA and...

  15. Plasmids 101: Codon usage bias

    Type
    Blog Post
    ...found in highly expressed genes (Emilsson and Kurland, 1990). Controlling gene expression through gene sequence...closely related yeast strains. Adaptation to changing conditions - Organisms often need to express genes at ...change which proteins are highly expressed and which are poorly expressed by producing and charging specific...necessarily mean the gene is going to express well. If you do get good expression, you should also functionally...suitability for your expression host before using them in your experiments. Expression of alternative tRNAs...Plasmids 101: protein expression Plasmids 101: E. coli strains for protein expression Plasmids 101 featured...interest, it’s possible to overexpress low abundance tRNAs in your expression host and thereby increase...

  16. To Codon Optimize or Not: That is the Question

    Type
    Blog Post
    ...Cells however, may not express all 61 of these tRNAs and those tRNAs that are expressed may be found at very...extremely useful in expressing functional proteins in hosts that do not naturally express that gene. The protein...from eukaryotic organisms for expression in prokaryotic systems or yeast (Lanza et al., 2014). For example...optimizing a human gene for expression in E. coli. Genes are also optimized for expression in mammalian cells.... for expression in mammalian cells. This was the first codon optimized membrane protein expressed in mammalian...this mean for a molecular biologist who wants to express genes from one organism in another? Let’s take ...and when you might want to optimize codons for expression in a particular organism. Why is codon usage ...

  17. A Practical Approach to Choosing the B(right)est Fluorescent Protein

    Type
    Blog Post
    ...variation, a reference fluorescent protein that is expressed at an equal level (Goedhart et al, 2011) can be...2012, Bindels et al, 2017). The plasmid for co-expressing mTurquoise2 with mVenus as a reference protein...brightness can be made. This strategy has been used in yeast by Lee et al (2013) and in nematodes by El Mouridi... The practical brightness assay (employing co-expressing fluorescent proteins or using endogenous tagged...

  18. RaPID Detection of RNA-protein Interactions

    Type
    Blog Post
    ...Run) and to detect protein-protein interactions (yeast-two hybrid, Co-IP, BioID),  and that’s just to name... study endogenous RNAs at their physiological expression levels; however, knock-in of BoxB sites into ...endogenous DNA sequences could allow one to study RNAs expressed from their native regulatory elements. Controls...evidence that suggests changes in RC2H1 protein expression and post-transcriptional regulation of SM1v1-...identified QKI, an RNA-binding protein that’s highly expressed in neural progenitor cells. Knock-down of QKI ...

  19. Some Like it Hot: Thermostable GeoCas9

    Type
    Blog Post
    ...for a number of model organisms: mammalian cells, yeast, drosophila, plants, worms, zebrafish, frogs, some...temperatures ranging from 25-80 °C. Activity is expressed as Kcleave (min-1). SpCas9 has maximum activity...

  20. Plasmids 101: Degron Tags

    Type
    Blog Post
    ...plasmid-based expression systems. Degron tags are one of many tools to manipulate protein expression. These ...the modulation of endogenously and exogenously expressed proteins. Here we will review what advantages ...used to manipulate endogenously and exogenously expressed proteins. Thirdly, degrons and their drugs are...kDa tag. Next, the TIR1 F-box protein must be expressed (inducibly, ectopically, stably… it’s your call... protein will associate with the endogenously expressed SCF complex members, an E3 ubiquitin ligase complex...requires an additional component to be engineered or expressed beyond the tag and small molecule. It does work...work in mammalian cells, mice, and even yeast, though (we have a kit!). Choosing a degron tag Are all tags...
Showing: 41 - 60 of 130 results