Endy: DNA sequencing

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How to submit a DNA sample for sequencing:

  1. Go to the Biopolymer lab website and read the page on DNA sequencing.
  2. Prepare your samples:
  • Samples should be prepared in 8-strip PCR tubes with lids.
  • Samples should not exceed a volume of 12µl; calculate ahead of time how much water, DNA, and primer you will add.
  • First, add the water. If using a custom primer (like VF2 or VR), add your primer (final amount should be 3.2pmol; use 2µl of a 1.6pmol/µl dilution of primer). Finally, add your DNA (see Biopolymer lab website for amounts; use 200-300ng of plasmid DNA).
  • Once everything has been added, close the tubes w/ a strip of caps.
  1. Go to dnalims.mit.edu and login.
  2. Go to Enter DNA sequencing requests.
  3. Input the number of samples and fill out the rest of the sample information. Use the Endylab account # for the order. Be careful not to mix up your samples. You want to be sure you know what sequences that you get back go with which samples.
  4. Submit the request and print two copies of the receipt. Keep one for Endylab records and take one with you to the sequencing facility.
  5. Bring your samples and printout to the sequencing facility (4th floor in the cancer center; enter by the entrance opposite the south end of bldg 68). Put your samples in one of the empty falcon tubes, label the tube w/ the order # and your name, and leave the tube in the fridge. Put your printout in the sleeve on the fridge.
  6. When you get the email saying your sequences have been completed, log into dnalims.mit.edu and go to retrieve sequences. Download the chromatogram and analyze it using ApE.
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