Protein Alignments and Phylogenies I

Inquiry-based Marine Biotechnology and Bioinformatics for Teachers
	What to do when you get your
	Sequences!

Sequences are typically returned from the sequencing facility as attachments to an email like this one.

We are going to pretend that you have received the email, saved the attachment to a folder and decompressed it using a utility such as stuffit (30-days free).

1. Click on "My Computer" and go to "Student on Atlantic/Ocean H:"

2. Then choose "temp2004"

then "0_Teacher_Enhan"

then "Ze_Sequencing_Results"

3. Now highlight that folder.

4. Go up to "Edit" and choose "Copy"

5. Paste the folder inside your folder... ie., double-click on your name, choose "Edit" and "Paste"

It might take a while because there are a ton of files inside the folder and you are all copying from the same hard drive.

After you have your own copy of this folder, have a look inside. Each of the two next folders represent the contents of a single email attachment from the sequencing facilty. They were uncompressed using Stuffit, and saved to this hard drive so that we could get at them quickly. Ordinarily you would receive the email, save the attachment, uncompress them, and save them to an appropriate folder.

1. Open Chromas

Download the zip file containing Chromas and save it in your folder called "programs." Go to the folder and let the zipfile self-extract. Then open Chromas. Chromas reads files with the ".abi" extension. You will understand the names of these files better than I do because they are your cloned pcr products!

Your job is now to open your sequences in Chromas and then export them to FASTA format so that you can open them in NotePad and clean up the sequences. You should use your lab notebook to help you decide what to name the sequences. WAIT for Simona to join us in the lab before editing your sequences...

Choose "Edit" ---> "Copy Sequence" ---> "FASTA Format"
Then go to "File" ---> "Export"
Check that the format is FASTA and give the file a name such as "Jetty_Californianus_seq.nuc"
Do this for all your sequences and then take a break... get up and walk around.
Using "Notepad," open one of your new FASTA files and notice the very important format.

Line 1: >The name and information goes here
Line 2: HERE IS THE SEQUENCE ONLY, NOTHING ELSE, ESPECIALLY NO EMPTY LINES

Use ctrl-f to find the first "N" and wait for Simona.

After you are done cleaning up the FASTA sequences, cut and paste one of them into the Blast Nucleotide-Nucleotide search window.

Hopefully your search will return a list of sequences of genes similar to the one you are looking for. If not, your next step will be to find the largest open reading frame, translate it and save the resulting amino acid sequence to a file in the FASTA format and submit that sequence to a BLAST protein-protein search... OR go back to the mussels and try again.