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Addgene

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

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We narrowed to 42 results for: ins

Showing: 21 - 40 of 42 results

  1. Gibson Assembly Protocol

    Type
    Protocol
    ... Gibson and colleagues at the J. Craig Venter Institute developed a novel method for the easy assembly...strong secondary structures in the homology region. Hairpins in this region can significantly reduce the efficiency...annealed DNA fragments. Taq DNA Ligase - covalently joins the annealed complementary DNA fragments, removing...for 1 hour at 50 °C or follow manufacturer's instructions. You can purchase master mix or make your own...into the assembly and is used when the part to be inserted is too long to include on overlapping PCR primers...

  2. Antibody Validation Using the Indirect ELISA Method

    Type
    Protocol
    ...protocol describes how to run an indirect ELISA against a purified antigen to demonstrate the expected ...indirect Enzyme-Linked ImmunoSorbent Assay (ELISA) against a purified antigen to validate an antibody. This... the antigen and how to probe with an antibody against the target to show a dose response. Sharing speeds...several days by incubating certain steps at 4 °C instead of room temperature or 37 °C. The protocol notes...minutes. Absorbance was read at 450 nm and plotted against the micrograms of antigen loaded. Last reviewed...

  3. What is Polymerase Chain Reaction (PCR)

    Type
    Protocol
    ...capabilities. Note: See manufacturer’s instructions for specific instructions about extension time and temperatures...reference page on how to design primers for detailed instructions and check out our protocol on plasmid cloning...each ingredient specifically do? Template DNA: Contains the portion of DNA that we are interested in amplifying...

  4. AAV ddPCR Titration

    Type
    Protocol
    ...depending on the insert. Please ensure that you are in compliance with your institution’s biosafety regulations... To ensure that the droplets are not disrupted, insert the pipette tips directly in the center of the ...Pro-Tip To ensure that the droplets are not disrupted insert the pipette tips and gently touch the bottom of...

  5. Kit Free RNA Extraction

    Type
    Protocol
    ...LiCl selectively precipitates RNA versus DNA or proteins. Add the correct amount of 7.5 M LiCl solution...downstream applications. To improve yield of RNA, instead of incubating at -20°C for 1 hour, you can try ...

  6. AAV Production in HEK293 Cells

    Type
    Protocol
    ...For each T-175 flask: Aspirate culture media and rinse once with 10 mL of PBS. Aspirate PBS and add 2 mL... at ~80% confluence. Aspirate culture media and rinse once with 60 mL PBS. Aspirate PBS and add 35 mL ... then transfer the cells into a sterile bottle. Rinse the CS2 with 100 mL of DMEM complete medium and ...Transfer cells and media into 500 mL conical vessels. Rinse CS5 once with 100 mL of PBS and add to the harvested...

  7. Plasmid Modification by Annealed Oligo Cloning (with Protocols)

    Type
    Protocol
    ...annealed oligos with cut vector in molar ratios (vector:insert) between 4:3 and 1:6 in a standard ligation ...ligation reaction (ex. to ligate an annealed oligo insert of 50bp in length into a 5kb vector, mix 100ng of the... multiple colonies for mini-prepping and verify insert by sequencing. Reference Page Top Index...

  8. Protocol - How to Inoculate a Bacterial Culture

    Type
    Protocol
    ...pre-mixed LB agar powder, use the suggested amount, instead of the other dry ingredients above. Media without...in a shaking incubator. Note: Some plasmids or strains require growth at 30°C. If so, you will likely ... to be in the log phase of growth. Check the instructions for your specific protocol and conduct an OD600...

  9. Protocols for Molecular Biology, Plasmid Cloning, and Viral Preps

    Type
    Protocol
    ...Creating Bacterial Glycerol Stocks Store bacterial strains or plasmids for long term use Watch the Video! ...Video! Western Blot Separate and detect specific proteins Watch the Video! Immunocytochemistry Use antibodies...the Indirect ELISA Method Run an indirect ELISA against a purified antigen to demonstrate the expected ...

  10. Using a Light Microscope Protocol

    Type
    Protocol
    ...of light hitting the sample or b) using dyes or stains that add color to some components of the sample..., be sure to record the magnification used, any stains used in your sample, and sample information so ...future. Conclusion Like any important laboratory instrument, you should be sure to take care of your microscope...

  11. Affinity Purification of Recombinant Antibodies with Protein A or Protein G

    Type
    Protocol
    ...should be properly disposed of following your institution’s hazardous waste procedures. Last Update: February...February 7, 2022 Protocol Video Watch this instructional video to learn how to affinity purify recombinant...for your columns, refer to the manufacturer’s instructions. Attach the Gravitrap column to a clamp on a...

  12. Protocol - How to Design Primers

    Type
    Protocol
    ... also use a nucleotide-based molecular insert (inosine) instead of a regular nucleotide for broader pairing...

  13. Protocol - How to Streak a Plate

    Type
    Protocol
    ...of plasmids in your purified DNA. This protocol explains how to isolate a single bacterial colony by streaking...plasmids or bacteria need to be grown at 30 °C instead of 37 °C. This is often true for large unstable...

  14. Lentivirus Production

    Type
    Protocol
    ...Envelope plasmid, and Transfer plasmid encoding your insert of interest Reagent Preparation DMEM Complete (... optimized for each transfer plasmid. Pro-Tip Endotoxins can inhibit transfection; therefore, plasmid ...

  15. Transfection for Recombinant Antibodies

    Type
    Protocol
    ...ensure that you are in compliance with your institution’s biosafety regulations. Last Update: February... °C. See below for other Reagent Preparation instructions. Wipe down all pipettes and equipment with 10%...

  16. AAV Purification by Iodixanol Gradient Ultracentrifugation

    Type
    Protocol
    ...purification and isolation of cells, organelles, lipoproteins, and macromolecules. Importantly, iodixanol ...of contaminants in each fraction. * AAV capsid proteins VP1, VP2, VP3; M protein marker. Image adapted...

  17. Isolating a Monoclonal Cell Population by Limiting Dilution

    Type
    Protocol
    ...cloning cylinders. Workflow Timeline This protocol begins with a stable cell pool. For details, see our protocol...homogenized cell solution to 990 µL complete medium. Instead of transferring 0.125 µL, now transfer 100 times...

  18. DNA Quantification

    Type
    Protocol
    ...you have a NanoDrop, follow the manufacturer's instructions for the spectrophotometer specific to your lab...

  20. Fluorescence Titering Assay

    Type
    Protocol
    ...ensure that you are in compliance with your institution’s biosafety regulations. Workflow Timeline Day...
Showing: 21 - 40 of 42 results