User:Anthony Salvagno/Notebook/Research/2009/10/15/Making Tethers

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Contents

Supplies

  • I made 1x popping buffer
  • I made k-casein in 1x pop (5mg/ml) hopefully this is a good blocker
  • Need a-dig
  • DNA
  • beads

A-dig

For this there are 20ul aliquots and I need 1:10 of that so add 180ul 1x Pop

DNA

I can try and tether both strectching DNA 4.4 kb and some unzipping stuff. I should dilute about 1:10 fold for both so that I can see if I got anything. I think we went more dilute, but I want tethers.

Beads

I need to dilute beads (amount TBD after looking through notebook), then sonicate to remove clumps. Following this protocol: User:Anthony Salvagno/Notebook/Research/2009/08/06/Bead Washing

OK Beads are now made. I don't think sonicating worked well.

Tethering

Ok, now everything is ready. It is time to tether.

  1. Flow a-dig
    • allow to sit 5min
  2. flow BGB
    • allow to sit 2min
  3. flow DNA
    • flowing 4.4kb stretch DNA and TD(1) unzipping DNA
    • allow to sit 5min
  4. wash with Pop
  5. flow Spheres
    • allow to sit 7min
  6. wash with pop
  7. Seal with nail polish
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