User:Anthony Salvagno/Notebook/Research/2010/05/17/Back in the habit, project planning for the immediate and future future

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No more shenanigans, it's time to get down to business! I'm back in the lab after a two week hiatus and ready to kick ass (to say the least). I have a lot to get done and I have 2 projects to take care of and so it looks like I have a bit of preplanning to do.

Project Lambda


So far I have:

  • performed the Klenow reaction
  • tested for adhesion
    • I'm pretty confident this worked because it looked like there was a shift in the band with the qd's. Of course I won't know for sure until one of two things happen:
      1. I do a reaction that fails
      2. We run the DNA through the microfluidics and that either works or fails.

To Do

The next step would be to take a little bit of DNA and adhere it to glass and attach qdots to the DNA. I'm not sure what the best way to go about this is. Here is Koch's opinion from an email conversation:

I think your best bet is to just try labeling with QDs. Also, probably re-run the original reaction, remembering to heat the DNA first. The gel shift and all will probably be too hard. With dilute QDs, you will look for "couplets" to prove that the DNA is labeled. Also you can label DNA with YOYO-1 and a different-colored QD to see if it's labeled.

I then said:

I was thinking about that, but how would I separate the DNA from unbound QDs? If I flow the DNA through, would washing clear unbound qds and not remove the DNA? I'm not sure how this would work without glass adhesion. Any thoughts?

The follow up:

S400 column perhaps? Not sure if QDs would be too big. Probably I guess. You could do a test where you run plain QDs through S400 and see how much it reduces their concentration (via counting on slide, or even just nanodrop maybe).

Ok so here is my project plan:

  1. Run S400 column of just QDs.
  2. Run nanodrop on qd's and see if I can determine concentration from this.
  3. If this works then proceed with DNA adhesion and purification.
  4. If it fails then I will try some regular glass to DNA sticking

Note: I just realized now that I can use the S-400 columns to purify after the Klenow reaction. This should be done since Phenol:Chloroform and EtOH precipitation is annoying, long, and not perfect.

DNA Unzipping and Tweezers


I have unzippable tetherable DNA. Pranav and I completed construction on the tweezers. We are in the midst of running tests and would like to begin calibration and to unzip DNA very soon. There is a lot to be done though.

To Do

I will really need to talk with Pranav about what to do for this.