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Addgene
Showing: 1 - 20 of 22 results
  1. Handling Plasmids from Addgene - Purifying Plasmid DNA

    Type
    Protocol
    ...20 min. Resuspend dry DNA with TE (10 mM Tris-HCl pH 8, 0.1 mM EDTA). Pro-Tip DNA resuspension can take...culture of bacteria transformed with your plasmid Resuspension buffer Denaturing solution Renaturing solution...several tubes/bottles. Remove the supernatant and resuspend the bacteria in buffer. Note: This step gets all... all of the bacteria back into suspension, but within a smaller volume of buffer that is compatible with...next solution. Add a denaturing solution to the resuspended bacteria. Note: This step causes the bacteria...column with 70% ethanol to remove excess salt. Resuspend the DNA pellet, or elute the DNA off of the column...Prepare the following solutions: Solution I - Resuspension Buffer 25 mM Tris-HCl (pH 8) 50 mM glucose 10...
  2. Pipetting Protocol

    Type
    Protocol
    ...release the liquid. Pro-Tip Dispense until the first stop, wait, and then dispense to the second stop to expel...selecting the correct pipettor and pipette tip, how to dispense liquids, and how to handle the pipette. Protocols... use a pipette in many scenarios to accurately dispense small amounts of liquid (think: 0.1 µL to 1 mL...mL). When working in a laboratory, properly dispensing liquid ensures the accuracy of your experiment ...experiment and any changes to the amount you are dispensing can negatively impact your experimental results. This...The table shows the pipette with the appropriate dispense volume: This image shows the P2, P10, P20, P100...and P1000 as shown from left to right. Pipette Dispense Volume P2 0.2 to 2 µL P10 1 to 10 µL P20 2 to ...
  3. AAV Production in HEK293 Cells

    Type
    Protocol
    ...Pipet back and forth to resuspend each pellet completely. Combine resuspended pellets and keep on ice....Pipet back and forth to resuspend each pellet completely. Combine all resuspended cell pellets and sonicate...containing the resuspended virus from step 12 above. Add 50 units of benzonase per mL of viral suspension (you ...cells after 30 passages. Procedure Trypsinize and resuspend the HEK293T cells from 2 x T-175 flasks. Cells...vigorously multiple times to obtain a single cell suspension (no clumps of cells). Pool cells from 2 x T-175... Return to incubator for 48 h. Trypsinize and resuspend cells from the CS2. Cells should be at ~80% confluence...forth or shake vigorously to obtain a single cell suspension (no clumps). Count cells using a hemocytometer...
  4. Kit Free RNA Extraction

    Type
    Protocol
    ...tissues, extraction of RNA, precipitation, and resuspension. This protocol provides two options: using Solution...RNA in the bottom of the tube. Wash the RNA by resuspending the pellet in 0.5–1 ml of 75% Ethanol and vortex...important to not let the pellet get too dry before resuspending, as this affects the solubility of the RNA. ... but while the white pellet is still visible. Resuspend RNA pellet in RNase-free water or TE. Quantify...
  5. Transfection for Recombinant Antibodies

    Type
    Protocol
    ...microcentrifuge tube. Vortex the cell suspension. Transfer 10 µL of cell suspension into the microfuge tube containing...culture. This protocol describes how to transfect suspension HEK293 cells with recombinant antibody plasmids... 5 mL pipette, transfer 0.5 mL of HEK293 cell suspension into a clean microcentrifuge tube. Pro-Tip Cells...Cells settle quickly and need to be resuspended before sampling. Gently swirl the flask 5–10 times before...Pipette 10 times to mix. Load 10 µL of the cell suspension/trypan blue mix into a cell counting chamber....
  6. Lentivirus ddPCR Titration

    Type
    Protocol
    ... media and 11.1 µg/mL Polybrene. Mix the cell suspension well before seeding. Mix each well with a 1 mL...by incubating with 200 µL TrypLE for 1–2 min. Resuspend cells in 500 µL DMEM complete and transfer to ...the master mix to each PCR tube. Be careful to dispense to the bottom of the tube without collecting drops...45° angle. Count to 20 while slowly and gently dispensing the droplets down the side of the tube. Place...
  7. Weighing Reagents Protocol

    Type
    Protocol
    ...of reagent that will be resuspended in a small volume (e.g. 0.02 g to resuspend in 1 mL), you may place...
  8. Western Blot

    Type
    Protocol
    ...5 min at 100 x g . Gently remove supernatant. Resuspend cell pellet in 1 mL of 1X PBS and transfer to ...min at 100 x g . Carefully remove supernatant. Resuspend cell pellet in appropriate volume of cold lysis...
  9. AAV ddPCR Titration

    Type
    Protocol
    ...the master mix to each PCR tube. Be careful to dispense to the bottom of the tube without collecting drops...45° angle. Count to 20 while slowly and gently dispensing the droplets down the side of the tube. Place...
  10. CRISPR Library Amplification

    Type
    Protocol
    ...for all plates. Use two 10 mL pipettes (one for dispensing cold LB and one for removing LB containing bacteria...cells are subject to lysis by freeze-thaw if not suspended in cryoprotectant like Glycerol or DMSO solutions...
  11. DNA Quantification

    Type
    Protocol
    ...spectrophotometer using the solution the DNA is resuspended in, but with no DNA added. 'Blanking' measures...
  12. Fluorescence Titering Assay

    Type
    Protocol
    ... pipetting or inverting. Aliquot 2 mL of cell suspension into each well of the 6-well dish. Incubate the...
  13. Colony Formation Titering Assay

    Type
    Protocol
    ...pipetting or inverting. Aliquot 1.35 mL of the cell suspension into each well of the 6-well dish directly on...
  14. Pouring LB Agar Plates

    Type
    Protocol
    ...during the autoclave process if your sample has spent at least 10 min at 121 ℃. Use lab tape to label ...
Showing: 1 - 20 of 22 results