TO SEQUENCE:

1. Submit an online sequencing request. Go to http://finch.daf.jhmi.edu
Type in user name and password.

2. Click New Request under Sequencing on the sidebar.

3. Select appropriate entries for DNA Type, and Bill To categories.

4. Click Tube Request further down the page.

5. Add all of your samples into the request by the following protocol.
-Click on a tube, so that it is highlighted in blue.

-Type in the template name and then the primer name in the
appropriate boxes.

-Click Set Selected button.

If you did it right, a double helix will appear in the picture of the tube.
The form automatically names each of your tubes as “TemplateName- PrimerName”.

You can not use single quotes in the sequence name (ex. 5'Not will not work, but 5Not will)

6. When finished adding all of your samples, click Add New Request button to submit your request.

Make sure that your request shows up under Pending! If it is not there, the request did not go through correctly.

7. Prepare samples to be sequenced. Pre-mix the primers and templates. Only 1 primer & 1 template per tube.

i.e. If you have 2 primers for one sample, set up two tubes, one with template + one primer and the other with the template and the other primer.

**Label your tubes exactly as you named the tubes in the online request order**

Amount to Send for Sequencing:

6uL of 0.2ug/uL template + 1uM primer

Since we have most of our primers in 20uM stock, it is easiest to send 20uL. that way you just add 1uL primer, X amount of template so you have 0.2ug/uL concentration, and Y amount of water to bring the volume to 20uL.

8. Bring samples to 1005 Blalock – Sequencing Facility. (Phone: 410-955-2836)