Question 3 in "A Simple Annotation Problem" handou

[cmackinnon]

I'm confused by the instructions on pg 4, question 3 on mapping each exon using BLAST searches. The instructions state: "Go to the NCBI BLAST search site and compare the protein sequence of the first coding exon of PMCA to the entire fosmid sequence." The last sentence states: Using blastx the results of this search look like this: [table shown, which I can't replicate]. How do I enter both the entire fosmid sequence and the sequence of exon 1 in a blastx window?

[cmackinnon]

Using Ruth Howe's notes, I see I need to check the box to align two or more sequences in the NCBI BLAST blastx choice. The query sequence is fosmid's DNA [obtained from the UCSC Genome Browser] and the subject sequence is the Gene Record Finder polypeptide details of the first exon. Let's see if I can do this again tomorrow!

[cshaffer]

Yes indeed,

you are correct. When comparing sequences with each other (in NCBI blast) you need to click that little check box "Align two or more sequences".

You will then get two input boxes one for each sequence.

Bonus material for advanced users: you can actually put more than one sequence in either box and NCBI blast will run all the searches at one time. Here is how the BLAST help explains it:

Align two or more sequences using BLAST. To use BLAST to align sequences one or more queries should be entered in the top text box and one or more subject sequences should be entered in the lower text box. Then use the BLAST button at the bottom of the page to align your sequences

This can save you time if you enter all the exon sequences in the bottom box and the fosmid sequence in the top box and do all the searches in one go.