Kun:LabNotes/ASE/2007-11-5

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Revision as of 23:48, 5 November 2007 by KunZhang (talk | contribs) (New page: ===Exp. 1=== Purify PCR amplicons with QiaQuick columns, 400ul reactions per column. <b>During the purification, I wasn't completely sure I added the amplicons to the tubes with correct la...)
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Exp. 1

Purify PCR amplicons with QiaQuick columns, 400ul reactions per column. During the purification, I wasn't completely sure I added the amplicons to the tubes with correct label. There is a possibility that I did it in the wrong order:

      Lib #1 -> tube #2     43ng/ul x ~100ul
      Lib #2 -> tube #3     63ng/ul x ~100ul
      Lib #3 -> tube #4     53ng/ul x ~100ul
      Lib #4 -> tube #5     54ng/ul x ~100ul
      Lib #5 -> tube #1     41ng/ul x ~100ul
      Lib #1 -> tube #1.2   86ng/ul x ~50ul

I decided to proceed with the purification. I purified another 400ul Lib #1 with a separate column.

I also performed ethanol precipitation on 400ul of Lib #2-5. In addition, I repeat PCR on Lib #1&2 using iTaq.

BciV digestion
To the ~45ul of column purified tube #1.2 and ~45ul ethanol precipitated Lib #2-5, add 5ul NEBuffer 4, 4ul BciVI, 37C 3h, keep @4C, check with 8% TBE gel.

   File:2007-11-05-BciVI-cut.jpg