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We narrowed to 1,051 results for: FER

Showing: 161 - 180 of 1051 results

  1. Quick Guide to All Things Lentivirus

    Type
    Blog Post
    ...stable expression of the transgene   Transfer plasmids The transfer plasmid is the vector you clone your...the control of two different promoters. Both the single and dual promoter transfer vectors can be used...genetically modify primary cells. Amongst the different vectors used, lentiviral vector constructs have...background Until the development of viral vectors, gene transfer into mammalian cells often relied on chemical ...These techniques are still used and efficiently transfer plasmid DNA efficiently into dividing immortalized...lines but reach their limits when it comes to transferring DNA into primary cells or non-dividing cells... particle (Gag, Pol, Tat, Rev and Env) and the transfer plasmid encoding the insert of interest as well...

  2. Antibody Guide

    Type
    Guide
    ... dates. Buffers Buffers are a concern for conjugating and signaling molecules, as some buffer components...heavy chain constant region” or Fc. Different isotypes play different roles in the immune response, and ...removed from the buffer before the reaction. However, it can be added back to buffers after the conjugation...of colors activated by different wavelengths, allowing for easy differentiation between multiple antibodies... In some cases, it may be preferable to buy small aliquots of many different antibodies and do batch validation...an antigen, while the specific binding site is referred to as an epitope. Production Antibodies are typically...typically produced in one of three different methods: in animals, in cultured immune cells, or through...

  3. Your Lentiviral Plasmid FAQs Answered

    Type
    Blog Post
    .... While these differences may seem subtle they result in a key physiological difference between lenti-...3rd-generation transfer vectors apart? A3: Addgene defines 2nd- or 3rd-generation transfer vectors based...packaging systems differ between 2nd- and 3rd-generation vectors? What does this mean for transfer vectors? A4...protein and consequently are considered safer. Second-generation transfer plasmids MUST be packaged with a 2nd-generation... Are transfer plasmids replication competent? What is SIN? A6: Most (if not all) Addgene transfer plasmids...which is rarely more than 30% of the transfer plasmid. Many transfer vectors are SIN. Packaging plasmid...ingredients: the lentiviral packaging vector, a transfer vector including the gene of interest, and the...

  4. Adeno-associated virus (AAV) Guide

    Type
    Guide
    ... via a different plasmid. In order to produce rAAV vectors, three plasmids are needed: Transfer plasmid...s Molecular Biology Reference . Production The three production plasmids (transfer, packaging, and helper...2, and 5 display differential efficiency and cell tropism after delivery to different regions of the central...immune response. These features have made AAVs the preferred choice to be used as viral vectors for gene therapy...transgene, gRNA, or shRNA of interest into the transfer plasmid can be done with most standard cloning...restriction enzyme, Gibson Assembly, or Gateway. Some transfer plasmids may have limited restriction sites or...recombination of unstable regions, like the ITRs found in transfer plasmids. This will ensure that the repeats will...

  5. Gamma-Retroviral Vector Guide

    Type
    Guide
    ...three different plasmids containing the transfer, envelope, or packaging components. The transfer plasmid...packaging genes. However, they are different viruses and require different isoforms of these packaging components...type. From an experimental standpoint, the main difference between lentiviruses and gamma-retroviruses is...gamma-retroviral vectors, you need three plasmids (Figure 2): Transfer plasmid — contains transgene, sgRNA, or shRNA ... gene, due to a wide range of infectivity for different cell types (known as tropism). Figure 2: Gamma-retroviral...transgene, gRNA, or shRNA of interest into the transfer plasmid can be done with most standard cloning...restriction enzyme, Gibson Assembly, or Gateway. Some transfer plasmids may have limited restriction sites or...

  6. Mobile-CRISPRi: Bringing CRISPRi to Diverse Bacteria

    Type
    Blog Post
    ...Mobile-CRISPRi: a modular approach for CRISPRi transfer Mobile-CRISPRi offers two advantages over existing bacterial...chromosomal copy of the RP4 transfer machinery mobilize these plasmids for transfer to recipient cells during...disease. However, the rate of transfer was variable. They measured transfer rates by comparing the number... P. mirabilis, had only ~1 transfer per 1,000,000 colonies. Higher transfer rates are better suited for...Mobile-CRISPRi plasmids at Addgene! References Auchtung, Jennifer M., et al. "Biology of ICEBs1, an integrative...them. For example, gene knockdown with CRISPR interference (CRISPRi) has been limited to lab-adapted bacteria...diverse bacteria species. Existing protocols can transfer CRISPRi into a single bacterial strain, such as...

  7. Antibody Validation for Flow Cytometry

    Type
    Blog Post
    ...approach is RNA interference (RNAi). The most common method is transfection of short interfering RNA (siRNA...For CD365, they examined two different antibody clones produced by different vendors. The specific epitope... panels, where different manufacturers may have the same clone available with different conjugates. Summary...commercial antibody manufacturers. References and resources References Ayoubi, R., Ryan, J., Biddle, M. ...include a discussion of the Human Leucocyte Differentiation Antigens (HCDM) workshop approach, which focuses...one size fits all. The approaches used will be different depending on the sample and target type. To start... For intracellular targets, we usually test 3 different fix/perms (methanol, PFA-saponin, PFA-triton)....

  8. 28 Hot Plasmid Technologies from 2015

    Type
    Blog Post
    ..., A mutant Renilla luciferase, catalyzes the oxidation of its substrate, luciferin, which releases energy... neomycin or luciferase) to create a final lentiviral vector containing up to 5 different elements. Lentiviral...bioluminescence resonance energy transfer (BRET), whereby photons emitted by a Renilla luciferase variant are used to...alternative to the widely used luciferase reporter gene plasmid pGL3 pGL3 luciferase reporter gene plasmids... issues with fluorescent pH sensors, a set of luciferase-fluorophore pH fusion reporters, pRSETb-pHlash...pHlash, that use Bioluminescence Resonance Energy Transfer (BRET) were developed by the Johnson Lab . Characteristics...energy as a photon. The photon energy is transferred to the acceptor fluorophore, a circularly permuted...

  9. Multiplexed Overexpression of Regulatory Factors (MORF) collection

    Type
    Blog Post
    ... directed differentiation To understand how each human TF can contribute to differentiation, the Zhang...atlas. Computational analysis was used to infer differentiation trajectories and identify both up- and down-regulated... scientist at [email protected]! References and Resources  References Joung J. et al., A transcription...patterns determine everything from stem cell differentiation fate to tumor suppression (and most everything...construct. Cells were cultured following a differentiation protocol before performing single-cell RNA ...embryonic stem cells. Transduced stem cells were differentiated and single cell RNA-seq was performed to map...    This atlas linked TF expression to differentiation outcomes as well as global transcriptional ...

  10. Plan Your Experiment

    Type
    Guide
    ...many different Cas proteins available, and researchers can adapt these experiments for different organisms...Desired Genetic Manipulation Different genetic manipulations require different CRISPR components. Selecting...editor variants often have different optimal editing windows. Be sure to refer to the literature associated...discrepancies with the reference sequence you used for gRNA design. Addgene offers many plasmids containing...Activation/interference Needs good primer design Western blot Knockouts Activation/interference Needs a ...Expression and Delivery Validate Genetic Edit References CRISPR ( C lustered R egularly I nterspaced S...have expanded the CRISPR toolbox to include many different types of edits, including point mutations, base...

  11. Plasmids 101: Codon usage bias

    Type
    Blog Post
    ...most organisms on Earth, but different organisms have different preferences for the codons they use to ... organisms have different codon usage biases? The reasons for varied codon preferences among organisms...poorly charged tRNAs, different regions of the protein may be translated at different rates. The ribosome...Organisms often need to express genes at different levels under different conditions. With varied codon usage...constraints have molded which codons are used preferentially in which organisms - organisms have codon usage...your organism of choice. Unfortunately, codon preferences make it so you cannot choose among the possible... evolutionary constraints that lead to these preferences and what can we do about them? Read on to find...

  12. The Golden Stopwatch Award: Recognizing Facilitators of Research

    Type
    Blog Post
    ...your TTO have a question? Refer them to Addgene’s Tech Transfer Team: techtransfer@addgene.org Congratulations...Addgene Help Center: Tech Transfer Technology Transfer Information Tech Transfer News: A Reintroduction ...Addgene account for Tech Transfer Offices. They’re packed with tech transfer resources (summaries, electronic...stats and strategies with tech transfer offices. Drop us a line at techtransfer@addgene.org. Albert Einstein...requests from Addgene MTA: material transfer agreement Technology Transfer: the exchange of intellectual property...addgene.org identifies the hold-up. Your Material Transfer Agreement is stuck in “pending.” Fear not, there... to make MTA turnaround faster with technology transfer offices (TTOs). The latest of these efforts is...

  13. Adenovirus Guide

    Type
    Guide
    ... (A to G). Different serotypes exhibit varying tissue tropisms, meaning they preferentially infect certain...from different species. Used to alter infectivity and tropism. pShuttle Class of shuttle/transfer vectors.... These genes are now supplied in trans , via different plasmids or the packaging cell line, which not...need two plasmids: Shuttle plasmid (also known as transfer plasmid) — containing the transgene of interest...recombination between the transfected adenoviral transfer plasmid and the E1 region integrated in the packaging...vectors Third-generation rAdV vectors are also referred to as "gutless" or high capacity AdV (HCAdV) vectors...transgene, gRNA, or shRNA of interest into the transfer plasmid can be done with most standard cloning...

  14. Five Popular Model Organisms

    Type
    Blog Post
    ...utilized mice to study how different mutations in leukemia impact different treatment regimens (Zuber ...example, the Stainier lab studied the proliferation and differentiation of β-cells in the the developing zebrafish...β-cells that originate in different regions of the pancreas and produce different levels of insulin (Hesselson...mouse (Mus musculus) Let’s start with the most preferred mammalian model organism of many researchers: ...injecting mice with retroviral vectors that harbored different mutations found in leukemia, the Lowe lab was ... gene expression in specific cells of the fly (Pfeiffer et al., 2010). Yeast (Saccharomyces cerevisiae...at a time. Zebrafish embryos are also laid and fertilized externally, allowing scientists to easily manipulate...

  15. Deep Dive: Statistical Tests (Comparisons)

    Type
    Blog Post
    ...test assesses the sum of differences between pairs). Samples arising from different subjects or populations...test assesses the difference of the group means). Unpaired t-tests can also be referred to as two-sample...significant differences between the means of your groups. However, it will only tell you if differences exist...samples are different from each other, or if every permutation of pairs within the five is different. To identify...groups to determine if they are statistically different. They are classic parametric tests and one of ... whether or not two samples are statistically different than one another, a two-tailed test should be ...control group. If you only care if one group is different in a certain direction (i.e., you just want to...

  16. Antibodies 101: Beyond Surface Labeling

    Type
    Blog Post
    ..., Addgene uses 'labeling' to refer to antibodies and 'staining' to refer to small molecule dyes. Indirect...first fixed and then permeabilized, using two different buffer solutions. Fixing causes protein cross-linking...nucleus, a different fix/perm kit is required (to be precise, the fixing solution is different while the...plot (Figure 4b). Through this method, differences in proliferation between cell populations can be visualized...divisions).   Figure 4: Proliferation dyes can visualize proliferative capacity and speed of cells...lineage markers, like many of the cluster of differentiation (CD) proteins found on immune cells. However...use 'labeling' and 'staining' interchangeably to refer to any technique that combines a marker with a signal...

  17. Starter guide to induced pluripotent stem cells (iPSCs) part 1:  A renaissance in regenerative medicine

    Type
    Blog Post
    ...maintaining the undifferentiated “stem” state and, Their potency or capability to differentiate into specialized...special types of cells that can develop or “differentiate” into more specialized cells with specific functions...function adequately [1]. Stem cells’ ability to differentiate into multiple cell types makes them useful models... cell has the ability to produce all of the differentiated cells in an organism, including the embryonic...Pluripotency: When a cell has the ability to differentiate into cells of any of the three germ layers: ...detail below [2]. Multipotency: When a cell can differentiate into multiple cell types of the same lineage...name suggests, cells under this category can differentiate into only one cell type. Muscle satellite cells...

  18. Tips for a 1st time AAV user (by a Rookie AAV user)

    Type
    Blog Post
    ... virus can be stored at 4 °C and buffer exchanged the next day. Refer to the AAV Iodixanol Gradient Ultracentrifugation... AAV packaging plasmids. The cells need three different plasmids to produce AAV: 1) the RepCap plasmid...help mediate AAV viral replication; and 3) the transfer plasmid which encodes a transgene of interest ...more details. Pro tips I’m not going to lie, AAV transfer plasmids are kind of a pain to work with. These...structures that are prone to deletion from the transfer plasmid. The good news is there’s a few simple...to grow multiple bacterial cultures of your AAV transfer plasmid at 30 °C instead of 37 °C and then screen...restriction sites). Another way is to transform AAV transfer plasmids into bacterial strains, like NEB Stable...

  19. Plasmids 101: Transformation, Transduction, Bacterial Conjugation, and Transfection

    Type
    Blog Post
    ...created phages. Transfer of this DNA to another cell is referred to as transduction. Transferred DNA once inside...donor can transfer genetic material to the recipient bacterium. The genetic material transferred is commonly...Horizontal gene transfer (HGT) is the movement of genetic material between organisms. It plays a key ...unique to the lab. Let’s take a look at these different methods of DNA insertion. Transformation Transformation...Various factors promote natural transformation in different bacteria such as growth phase of the cells (Baltrus...plasmid, the genes on the plasmid can give them different phenotypes. For example, this plasmid encodes ...cell. Once inside, phages can follow one of two different life cycles: lytic or lysogenic. Lytic phages ...

  20. Viral Vectors 101: Gamma-Retroviral Packaging Systems

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    Blog Post
    ...pseudotype γ-retrovirus using different envelopes to modify tropism. Briefly, different envelope plasmids can ...engineered γ-retroviruses differ from lentiviruses? There are a few important differences between engineered ...packaging; however, they are different viruses and thus use slightly different isoforms of these packaging...Vectors 101 eBook, 1st edition.  References and resources References Swift S, Lorens J, Achacoso P, Nolan...following components are needed: γ-Retroviral transfer plasmid encoding a transgene, sgRNA, or shRNA ...sequences, which facilitate integration of the transfer plasmid sequences into the host genome. Typically... the host genome upon viral transduction. Most transfer plasmids available from Addgene were derived from...
Showing: 161 - 180 of 1051 results