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We narrowed to 902 results for: nes

Showing: 201 - 220 of 902 results

  1. Your Lentiviral Plasmid FAQs Answered

    Type
    Blog Post
    ...transgene but also that of adjacent cellular genes. Q7: Which genes are deleted or modified to result in replication...1]; and 2) the potential for oncogenesis through insertional mutagenesis. These risks are largely based...lentiviruses and gamma-retroviruses use the same genes for packaging (that's gag, pol, and env), the isoforms...specifically for transient expression (find some useful backbones here) or from viral transduction. While not ideal...cell line is used; several common laboratory cell lines including 293 were immortalized with adenoviral ...3rd-generation transfer vectors apart? A3: Addgene defines 2nd- or 3rd-generation transfer vectors based on...expression of lentiviral vectors in producer cell lines. Identifying less toxic envelope proteins or envelope...

  2. Controlling for Off-target Effects with a New Genome-wide CRISPR Screen Design

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    Blog Post
    ... identified ricin toxicity genes. The screen also discovered several genes previously not associated with...-throughput systematic approach for identifying genes involved in a biological process. These screens ...per gene and targets all ~20,500 protein-coding genes. Unique, non-overlapping sites in the genome were... pooled library via a lentiviral vector to cell lines that either stably expressed Cas9 or that had been...toxicity screen since essential and non-essential genes for both pathways had previously been identified...identifying >88% of previously identified essential genes while previous Cas9 and shRNA library screens only...only identified  60% of the essential genes. The ricin toxicity screen had a 10% false discovery rate ...

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

    Type
    Blog Post
    ...multiple genes with Gal4/UAS system One can also use Gal4 to drive expression of multiple genes under the... Drosophila geneticist’s main workhorse to turn genes on or off. Gal4 is a transcriptional activator that...machinery to the site to induce gene expression. Thus, genes and siRNA encoded downstream of the UAS sequence...promoter, and Bloomington’s Stock Center has fly lines expressing Gal4 under a variety of promoters resulting...quickly mate your UAS-YFG line with a variety of Gal4 lines and study gene expression in many different tissue...sequences. For example, if you discovered five new genes and wanted to express them within a specific neuronal...dosage effects. For example, overexpression of two genes by one Gal4 driver will produce roughly half as ...

  4. Fluorescent Proteins 101: Luciferase

    Type
    Blog Post
    ...disadvantages of luminescence versus fluorescence.  Table 1: Properties of Luminescence versus Fluorescence... the emission of photons. The most familiar bioluminescent organism for most people is the firefly (Photinus...surprisingly it is also the most commonly used bioluminescent reporter. This beetle emits a yellow-green ...as substrates. Production of photons by this bioluminescent reporter occurs slower than fluorescent-based...the different mechanisms to produce photons, chemiluminescent reporters are generally less bright than fluorescent...Fluorescence   Luminescence Fluorescence Source of Emitted Photons Chemical reaction High-energy photons...Photinus pyralis) is the most frequently used bioluminescent reporter, other luciferases have been identified...

  5. Five Popular Model Organisms

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    Blog Post
    ... They share about 70% of their genes with humans and 85% of human genes associated with a disease have...Zuber, Johannes, et al. "Mouse models of human AML accurately predict chemotherapy response." Genes & development...researchers around the globe. These organisms share many genes with humans, are easily maintained in the lab, and...manipulated with tools like CRISPR to make transgenic lines. The fruit fly (Drosophila melanogaster) Another... of Drosophila research. Morgan discovered that genes were found within chromosomes using the fruit fly... as it is estimated that 75% of disease causing genes in humans have a functional homolog in the fly (...packaged into chromosomes and about 23% of yeast genes have a counterpart in humans (Liu et al., 2017) ...

  6. Plasmids 101: The Wide World of Natural Plasmids

    Type
    Blog Post
    ...carrying genes for bacterial toxins like colicin typically also carry the necessary resistance genes to help...cell to another). Bacteria need special fertility genes to be capable of conjugation, and these are encoded...organisms to thrive in competitive environments. The genes encoding these functions form the basis of many ... Resistance (R) plasmids: microbial defense Genes allowing bacteria to survive toxins are the clearest...Nevertheless, the transfer of plasmids carrying these genes between bacteria can also have serious implications...: microbial infection Virulence plasmids encode genes that allow the host to infect other organisms. If...difference is virulence plasmids! This category includes genes that help bacteria invade hosts, including attaching...

  7. A New Generation of Adenine Base Editors Improves Editing in Primary Human Cells

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    Blog Post
    ...replaced S. pyogenes Cas9 in the ABE complex with PAM-variant Cas9 proteins: the S. pyogenes NG-Cas9 (PAM...While editing single genes is very useful, sometimes, you need to modify multiple genes to generate a cellular...adenine base editors with improved editing in cell lines Nicole Gaudelli, the Associate Director and Head...ABE8 constructs using a catalytically “dead” S. pyogenes Cas9 (dC9-ABE8.x-m/d). The dC9-ABE8.x-m/d constructs...adenine base editors The most commonly used S. pyogenes Cas9 requires the target site in the genomic DNA...(“N” can be any nucleotide base, followed by 2 guanines). This limits the use of ABEs as DNA sites that...In addition to stem cells the authors targeted 6 genes in primary human T cells using 8 selected ABE8s ...

  8. A Needle in a Base-Stack: Cas9 Structural Biology

    Type
    Blog Post
    ...from Streptococcus pyogenes.   Figure 2:  Crystal structure of S. pyogenes Cas9 in the apo state... we can see why: two arginines make base-specific hydrogen bonds to the guanines, but the N (a thymine...    Figure 3: Crystal structure of S. pyogenes Cas9 bound to guide RNA (PDB ID 4ZT0), shown opaque...the protein, spanning both lobes. Stem loop 1, nestled between the REC and CTD domains, is required for...protein portion of the CTD, ready to check for PAM sequencines. This helps explain why Cas9 can’t locate PAM...sequence isn’t in place yet.     Figure 4: S. pyogenes Cas9 in the apo state (PDB ID 4CMP, top) compared... residues in the CTD can get cozy with DNA. S. pyogenes Cas9 recognizes the PAM sequence 5’-NGG-3’, where...

  9. Supporting Rare Disease Research with BeHeard 2017

    Type
    Blog Post
    ...with other genes. He will use fluorescent proteins to monitor the localization of these genes, CRISPR to...to identify candidate genes as drivers of peritoneal mesothelioma tumorigenesis. They plan to use CRISPR...derived peritoneal mesothelioma cells lines and assess their tumorigenesis. These experiments will serve to...validate these candidate genes and allow Nabavi and colleagues to prioritize which genes they will use to develop... Di Schiavi will be able to better choose which genes and pathways to target in future small molecule ...molecules targeting SMN1 and its partners in mouse cell lines. Dr. Di Schiavi hopes that these studies will accelerate... research funding opportunities. References 1. Jones, C., et al. "PP09. 1–2352: Systematic review of ...

  10. Genome-wide Screening Using CRISPR

    Type
    Blog Post
    ...This post was updated on Aug 20, 2020. What genes are important in your phenotype of interest? Many scientists... cause is not entirely known. Identifying which genes are important for a phenotype can lead to a wealth...experiments investigating the role of individual genes or entire pathways in a particular disease process... can also be used to activate or repress target genes without permanently modifying the genome. If you'd... is to use Cas9 and a pool of gRNAs to identify genes that are essential for a given phenotype. Image ...screen a population of mutant cells to identify genes involved in a particular phenotype. CRISPR can be...created to knock out, activate or repress target genes by combining a gRNA library with the aforementioned...

  11. History of CRISPR Cas - A tale of survival and evolution

    Type
    Blog Post
    ...interspaced palindromic repeats) Cas (CRISPR associated genes) system. This popular genome engineering tool offers... be incorporated into the bacterial genome. Cas genes, which code for polymerases, nucleases, and helicases...adaptive immune system that utilized various cas genes not only to store a record of invading phages but...CRISPR/Cas is the system that scientists have harnessed for genome engineering. The diversity of the CRISPR... CRISPR/Cas systems provides powerful lines of defense against invading phages. Rising to the challenge...challenge - Evolution of anti-CRISPR genes in phages The CRISPR/Cas adaptive immune system seems like the winning...nucleases from re-identifying it (3). Surprisingly, few genes have been identified that neutralize CRISPR/Cas....

  12. An Introduction to Adenovirus

    Type
    Blog Post
    ...with the E1 and E3 (Early 1 and Early 3) genes deleted. These genes’ functions are rescued by helper plasmids...will be expressed, since the early genes are absent, and the late genes (and their structural proteins, ...the world, hypotheses began to develop that this illness was caused by either SARS-CoV-2, Adenovirus type... rarely cause severe disease in humans (and no vaccines are available on the commercial market). Although.... Its genome consists of “early” (E) and “late” genes, and the transition between early and late infection...replication (Harrach, 2014). Given that the early genes encode proteins essential to initiating the whole...plasmids used during production and/or by genes present in the HEK293T cell line used (which have fragments...

  13. New Optimized Genome-wide CRISPRko, CRISPRi, and CRISPRa Libraries

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    Blog Post
    ...of a gold standard set of genes (1,580 essential and 927 non-essential genes). The Root and Doench lab...viability. In addition, high copy number genes may mimic viability genes as they are subject to an increase ...help reveal the function of lowly expressed genes or genes that are more drastically affected by gene ...overexpression library identified several overlapping genes, most genes were unique to the individual libraries. These...be used to knock out, inhibit or activate target genes by combining specific sgRNAs with Cas9 or Cas9 derivatives... CRISPR libraries in the depletion of essential genes. In the subsampling analysis of the sgRNAs, Brunello...discriminate between non-essential and essential genes similarly to the Brunello knock-out library while...

  14. Minigenomes - a Safe Way to Study Dangerous Viruses Like the Ebola Virus

    Type
    Blog Post
    ...that this system uses plasmids where the genes (either viral genes or minigenome) are under control of a ...transcription (6). The EBOV genome contains seven viral genes, each flanked by a transcription start signal sequence...replication and transcription (Figure 1). All viral genes are removed and replaced by a single (non-viral)...replicates the viral genome and transcribes viral genes. Four viral proteins are required for viral genome...When used to study minigenomes, these four viral genes are each encoded on separate plasmids and are co-transfected...alongside a plasmid encoding the minigenome. These genes and the minigenome are transcribed (and the viral...is technically possible to encode all four viral genes in the same plasmid (as opposed to four separate...

  15. Quick Guide to All Things Lentivirus

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    Blog Post
    ...harmful genes into a transfer vector. Lentiviral particles expressing proto-oncogenes or oncogenes for instance...DNA efficiently into dividing immortalized cells lines but reach their limits when it comes to transferring...packaging plasmid containing all the structural genes necessary for the production of a viral particle...the packaging plasmids lacked the psi signal, the genes encoding the structural proteins could not be integrated...plasmid), 2) the HIV-1 genome containing all viral genes with the exception of the env gene (packaging plasmid...taken especially if you are working with proto-oncogenes or with human samples that have not been tested...generation because this vector requires only three HIV-1 genes (gag, pol, and rev) for production. Third generation...

  16. Advice for Moving Into Sales After Your Science Postdoc

    Type
    Blog Post
    ...studying cell division. However, after 6 years in the business of western blots and cloning, I decided to pack...with little emphasis placed on the marketing or business background of the applicant. However, although...push not only my academic background but also my business acumen and sales experience. Most scientists think... I tore up my “Scientific CV” and wrote a more business friendly CV that would focus on my achievements...conferences I presented at and placed them under headlines entitled “Presentation Experience”, “Career Achievements... profile, matched it with my CV and added more business-like elements. At this point you may be asking... the focus of your CV from  science-focused to business-focused?" The reason is that I knew that the mind-set...

  17. Hot Plasmids: Fall 2024

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    Blog Post
    ...CRISPR/Cas9 for gene disruption in M. abscessus  Bioluminescent reporter of mechanical forces   Epitope tag...expression levels comparable to GFP during early embryogenesis (Figure 3). mScarlet-I3 also offered clearer...development. Representative images of single focal planes through the middle of each embryo are shown, comparing...elegans: mScarlet3 for applications needing intense brightness (such as low-expression proteins of interest)...Here, the researchers were able to target multiple genes in M. abscessus either simultaneously or stepwise...e0033523. https://doi.org/10.1128/jb.00335-23.  Bioluminescent reporting of mechanical forces with PILATeS...expressed separately) assemble and catalyze the luminescence reaction (Figure 5). The researchers used the...

  18. Why and how to start studying chromatin (with Addgene's help)

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    Blog Post
    ...plasmids containing these genes. As with the PIs above, many of these genes were discussed at the 2018...context of chromatin?” Though it’s true that adding histones and chromatin architecture to every diagram portraying...off point. The following lists of PIs, articles, genes, and blog posts are not comprehensive, so just because...Zhang Leonard Zon  You!     Chromatin-related genes with plasmids available from Addgene The links below...Keystone meeting for Chromatin. Note that histone genes frequently have many different variant names, not...our website. In-exhaustive Chromatin-related genes with plasmids on Addgene.org AGO1 BRCA1 CREBBP... RBX1 STAG2 Xist Your favorite gene! SWI/SNF Genes:       BRG1(SMARCA4) BAF170 (SMARCC2) ARID1A...

  19. Stabilized Bacterial Promoters: Constant Gene Expression at any Copy Number

    Type
    Blog Post
    ...tested in different plasmid backbones that varied in size or carried different genes. In these cases, gene ...three genes whose proteins are needed sequentially. When Segall-Shapiro et al. placed the genes under ...Researchers express genes of interest from plasmids in order to study gene function or to engineer cells... fluctuations due to copy number by introducing genes of interest into the chromosome. But this has its...replication more than once per cell division and thus, genes closer to the origin of replication will be overrepresented...initiated before the cell has divided. This means that genes located near the origin will be over-represented...stabilized promoters were equivalent whether the genes were expressed from the genome or a plasmid. Thus...

  20. Technique: Probe Phage Genomes for Host Binding Proteins

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    Blog Post
    ...nitrocellulose membranes and exposed to IPTG in order to induce protein expression. Cells on the membranes will ...high salt.  To probe membranes with target bacterial cells, wash your membranes with PBST to remove salt...done, this technique can also help pinpoint the genes within a sequenced genome that encode for functional... vector. In order to reduce the chances of some clones being outcompeted by others in liquid culture, ...When your protein induction is done, peel the membranes off the plates and place each onto a lysis solution-soaked... gene encodes that protein! First, block your membranes with BSA or skim milk as you would for a Western...to remove cell debris. Then, UV-irradiate the membranes for 15 min to kill any remaining live E. coli....
Showing: 201 - 220 of 902 results