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Enter a sequence to perform a BLAST-based search of Addgene plasmid sequence databases.

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We narrowed to 974 results for: LAT

Showing: 941 - 960 of 974 results

  1. Colony Formation Titering Assay

    Type
    Protocol
    ...(μL) Volume of Virus Added to Plate (μL) Volume of Cells Added to Plate (μL) Final Viral Dilution 1:10...solution and, therefore, the cells cannot be used for later experiments. Workflow Timeline Day 0: Seed and transduce...integration events decreases as the dilution increases. Calculate the transduction units per mL (TU/mL) as follows...

  2. Handling Plasmids from Addgene - Purifying Plasmid DNA

    Type
    Protocol
    ... Gel DNA Purification Streaking and Isolating Bacteria Inoculating a Liquid Bacterial Culture Introduction...bacterial culture. For details on how to streak a plate to get individual colonies and to generate liquid... opens in a new window) Promega sell kits for isolating plasmid DNA in quantities as low as a few micrograms...

  3. Kit Free RNA Extraction

    Type
    Protocol
    ... Work in a well-ventilated space and under a fume hood when working with the volatile reagents in the ...nucleoprotein complexes. The effectiveness of your RNA isolation will depend on how effective your cell lysis protocol...

  4. AAV Production in HEK293 Cells

    Type
    Protocol
    ...controller Hazardous waste container pH meter Stir plate Magnetic stir bar Sonicator Ear protection Vortex...reach 100% confluence (80–90% is ideal). Pass and plate the cells the day before the transfection. Thaw ...incubator for 24 h–36 h. Proceed with transfection: Calculate the amount of each plasmid needed to have a 1:... Therefore, for each plasmid we need: Sample Calculations RepCap: 0.08 μg/bp × 7,265 bp = 582.1 μg Volume...

  5. Protocol - How to Create a Bacterial Glycerol Stock

    Type
    Protocol
    ...competent cells and retransform. Bacteria on an LB agar plate can be stored at 4°C for a few weeks. However, if...glycerol stocks. Procedure Follow the steps for Inoculating an Overnight Liquid Culture . After you have ...stock unthaw! Streak the bacteria onto an LB agar plate . Grow your bacteria overnight at the appropriate...

  6. Centrifugation

    Type
    Protocol
    ... the two, use a conversion calculator or chart like the G Force calculator (Link opens in a new window...protocol. Pro-Tip Spin speed is often given as Relative Centrifugal Force (RCF) or Revolutions Per Minute...

  7. Molecular Biology Protocol - Restriction Digest of Plasmid DNA

    Type
    Protocol
    ...reaction in 1 hour. Using this ratio, you can calculate the minimal amount of enzyme for your reaction...did not cut, check to make sure that it isn't methylation sensitive . Plasmids grown in Dam... (Link opens in a new window) NEB's table of methylation sensitive restriction sites . Sometimes enzymes...

  8. CRISPR Guide

    Type
    Guide
    ...genomic edits by copying from a DNA donor template . The donor template contains the desired sequence and is...linear dsDNA templates work better for large edits or insertions. When designing the donor template, incorporating...modifying the methylation state of cytosines in a gene’s promoter or by inducing histone acetylation or demethylation...through increased H3K27 acetylation TET1 - activation through cytosine demethylation DNMT3A - repression ...repression through cytosine methylation MQ1 - repression through cytosine methylation with improved kinetics LSD1...detection platform. Based on this non-specific cleavage, a Cas13a-based molecular detection platform, termed...pathway. HDR relies on copying DNA from a matching template to accurately repair or fill in the missing sequence...

  9. Plan Your Experiment

    Type
    Guide
    ...system that enables researchers to manipulate the genome with relative ease. Over the past decade, researchers... Select Your Desired Genetic Manipulation Different genetic manipulations require different CRISPR components... donor template Cas9 or Cas9 fusions with proteins that promote HDR Requires a repair template Lower editing...Does the resulting population need to be identical (clonal) or is a mixed population okay? Does the edit...modifications . Cell Population The first step of validation is determining the type of cell population you need ...you will likely need a clonal population. Additionally, clonal populations are useful for determining if...edited alleles. A clonal population is not always required, and a mixed population can be sufficient for ...

  10. Molecular Biology Reference

    Type
    Guide
    ...enable isolation and purification of cellular components, such as DNA, but practically this isolation is ...is only feasible for relatively short DNA molecules. In order to isolate a particular gene from human chromosomal... and manipulating genes. For instance, most plasmids are replicated in E. coli and are relatively small...20,000 basepairs) for physical isolation (purification) and manipulation. With current cloning technology...most commonly used is the Illumina NGS platform. This is the platform used by Addgene’s sequencing partner...of DNA, or templates, are spatially separated from each other. These attached DNA templates are then amplified... again producing ~1,000 copies of each template. Each template is then replicated using the modified bases...

  11. Antibody Guide

    Type
    Guide
    ...produces a colored product that accumulates over time which is relative to the amount of enzyme present...quantify relative amounts of protein. Includes: Western blot ELISA Capture - used to isolate specific ... one type of antibody. Monocultures of B cells isolated from animals immunized as described above are ... regions of the antibody to be independently manipulated. Antibodies produced this way are called recombinant... used while suspended in the original media or isolated and placed in a buffer. Antibody Structure Figure...antibody fragments (shown in Figure 2) to completely unrelated molecules. Examples include: scFv - Single-chain...instead of modified antibodies and are not closely related to antibodies. Monobodies - Monobodies are synthetic...

  12. Adenovirus Guide

    Type
    Guide
    ...necessary for viral DNA replication and host cell manipulation. Late transcripts are expressed after DNA replication...replication and host cell manipulation. L1, L2, L3, L4, L5 in trans Adenoviral late transcript regions that...are divided into early (E1A, E1B, E2, E3, E4) and late (L1, L2, L3, L4, L5) regions. Early transcripts ...also made them the most widely used viral vector platform for vaccine design against a diversity of viruses...which is how they got their name, but they are not related. AAVs require the presence of adenoviral genes ...and have been produced by replacing the early and late adenoviral transcript regions between the two ITRs... inside of the adenoviral capsid and directly correlates to the size of the DNA removed from the viral...

  13. Adeno-associated virus (AAV) Guide

    Type
    Guide
    ...hepatitis virus post‐transcriptional regulatory element; stimulates the expression of transgenes via increased...cis The polyadenylation or poly(A) signal aids in the nuclear export of RNA and RNA translation, and promotes...encodes for virus-associated RNAs involved in regulating translation and inhibiting cellular anti-viral defenses...which is how they got their name, but they are not related. AAVs require the presence of adenoviral genes ...available inside of the small AAV capsid and directly correlates to the size of the DNA removed from the viral...kidney 293T (HEK293T) cells are the most common platform to produce AAV vectors. Additional production ...cannot enter the lytic cycle, and instead establish latent infections by integrating into the host genome ...

  14. Chemogenetics Guide

    Type
    Guide
    ...previously used in human populations in Japan, making it an attractive option for translational studies, however...receptors, or ion channels targeted to specific cell populations (most often neurons) that can be turned on or...therefore allowing precise control over the cell population they are targeted to. To find chemogenetics plasmids...ligand. Ligand binding then activates G-proteins to modulate downstream signaling. GPCRs have been shown to...G-protein and activate neuronal firing through stimulating phospholipase C, which releases intracellular... and inhibit neuronal signaling by inhibiting adenylate cyclase and downstream cAMP production. There ...hM4Di, and is especially attractive for use in translational studies because it is FDA and EMA approved. ...

  15. Promoters

    Type
    Guide
    ...by various DNA regulatory sequences including enhancers, boundary elements, insulators, and silencers....RNA responsible for providing a stable template for the translation of a protein. RNA polymerase III transcribes...polymerase transcribes DNA to mRNA which is ultimately translated into a functional protein. Thus the promoter ...produced. The antisense strand is referred to as the template strand or non-coding strand as this is the strand...proteins which recruit RNA polymerase and control and regulate the transcription of DNA into mRNA. Promoter binding...promoter region stably and transcription of the template strand can initiate. The TATA box is a DNA sequence...proximal promoter which contains many primary regulatory elements. The proximal promoter is found approximately...

  16. Optogenetics Guide

    Type
    Guide
    ...silence the same neuronal population, or activate different neural populations at distinct times. Temporal... Two factors determine which neuronal population is manipulated in a given experiment: the expression ...optics and genetic engineering to measure and manipulate cells (frequently neurons) and their governing...neurons upon exposure to light through direct stimulation of ion channels. Naturally occurring channelrhodopsins... nonspecific cation influx into the cell when stimulated. Channelrhodopsin-2 (ChR2), the first widely ...Halorhodopsins are light-gated inward chloride pumps isolated from halobacteria. Wild-type halorhodopsin, known...High-efficiency blue-shifted channelrhodopsin from Platymonas subcordiformis 445 CoChR Channelrhodopsin from...

  17. Lentiviral Vector Guide

    Type
    Guide
    ...hepatitis virus post‐transcriptional regulatory element; stimulates the expression of transgenes via increased...single strand of RNA contains packaging genes, regulatory genes, accessory genes, and the long-terminal...Wild-type lentiviruses additionally require the regulatory genes tat and rev , along with virus-specific... — contains packaging genes gag and pol , and regulatory genes tat and rev ; separated into two plasmids...plasmid — contains entire viral genome (packaging, regulatory, and accessory genes), only envelope removed ...can more specifically dictate the target cell population and reduce cellular toxicity. Common Uses of ...(FACS) is used to sort cells expressing GFP and later expanded into a cell line. More information on generating...

  18. DNA Quantification

    Type
    Protocol
    ...Calculate your DNA sample concentration and purity using Addgene’s DNA Quantification Protocol....Quantification DNA Quantification You may also like... Inoculating a Liquid Bacterial Culture Recovering Plasmid...

  20. Water Bath Protocol

    Type
    Protocol
    ... water, not tap water, to prevent salts from accumulating on the surfaces of the water bath as the water... your experiment. Water baths tend to heat up relatively slowly. Place the water bath cover on the top...
Showing: 941 - 960 of 974 results