1. © Wiley Publishing. 2007. All Rights Reserved. How Most People Use Bioinformatics

2. Learning Objectives Get an overview of the most basic tools used in bioinformatics Get an overview of Medline, the virtual library Get an idea of what the rest of the course will be about

3. Outline Retrieving scientific information with Medline Fetching the protein or the DNA sequence you need Searching a database with BLAST Making a ClustalW alignment

4. PubMed/Medline PubMed is a database containing all the recent scientific publications in biology PubMed is free You can search PubMed using any keyword you are interested in.

5. Searching PubMed Rapidly Open www.ncbi.nlm.nih.gov/pubmed www.ncbi.nlm.nih.gov/pubmed Type your favorite keywords Press Return or Enter

6. Searching PubMed Precisely Click the Limits tab Check the boxes you are interested in, such as Review English AIDS

7. Searching PubMed Very Precisely Restrict the search with fields [AU] Author [SO] Source (journal) [TI] Title [AD]Address [MH]Keywords The words will be searched only in the corresponding fields

8. Tips for Searching Medline Use OR and AND to refine your queries Add the initials of the paper’s author, as in Smith TF Save the PMID of your papers Very precise Very short Medline contains only papers published after 1965 Use no more than 10 names for papers before 1995

9. Retrieving Protein Sequences in Swiss-Prot Swiss-Prot is a database containing all the proteins with known functions Swiss-Prot is available from the ExPAsy server at www.expasy.ch/sprot/ www.expasy.ch/sprot/ ExPASy: Expert Protein Analysis System ExPASy contains many useful online tools

10. The Swiss-Prot Entry Each Swiss-Prot entry is dedicated to a protein A Swiss-Prot entry summarizes everything that is known about a given protein The entry contains functional information and links to other databases mentioning this protein

11. Typical Swiss-Prot Entry Protein name Protein function Bibliography Links to other databases Structure Domains Function

12. Looking for DNA Sequences There are many types of DNA sequences The most common are Regulatory regions, often before genes Untranslated regions, often around the genes Protein-coding regions Intergenic regions (between the genes) All these sequences can be found in GenBank

13. Fetching a DNA Sequence at the NCBI Navigate to www.ncbi.nlm.nih.gov/Genbank/ www.ncbi.nlm.nih.gov/Genbank/ Type in a keyword. Press Return or Enter. You get a list of entries matching your keyword. Point, click, and explore…

14. Searching for Your Sequence with BLAST BLAST: Basic Local Alignment Search Tools Compares your sequence with all other sequences in your favorite database Returns the most similar sequence

15. Choosing Your BLAST Program Navigate to the NCBI BLAST site at www.ncbi.nlm.nih/BLAST Select your BLAST program: Blastn DNA BlastpProtein

16. Blasting a Protein Sequence Cut and paste your sequence Click the BLAST button at the bottom of the screen Wait

17. Reading a BLAST Output Every line is a hit Best hits come first Low E-Value = good hit E-value >1 = bad hit G means a link onto a complete genome U means a link to UniGene, the transcript database

18. Multiple-Sequence Alignments (MSAs) Multiple alignments reveal common features between sequences Multiple alignments are useful for Comparing very different sequences Making phylogenetic trees Making structure predictions Multiple-sequence alignments are abbreviated as MSAs

19. Making a Multiple-Sequence Alignment Identify a set of related sequences Do a BLAST of your favorite sequence Choose a method: www.ebi.ac.uk/clustalwClustalWPopularwww.ebi.ac.uk/clustalw www.tcoffee.orgT-CoffeeAccuratewww.tcoffee.org www.drive5.com/muscle/MuscleFastwww.drive5.com/muscle/ www.tcoffee.orgM-CoffeeConsensuswww.tcoffee.org

20. Making an MSA with M-Coffee Open www.tcoffee.org Click MCoffee::Regular Cut and paste your sequences Submit your MSA

21. Making Sense of Your MSA Positions are marked: Completely conserved = asterisk ( * ) Highly conserved = colon (:) Conserved = period (.) Look for highly conserved blocks: The red box on this slide shows a highly conserved block. These blocks are often functionally important positions.

22. Going Farther Bioinformatics is all about getting knowledge without having to make real-world experiments. Many more details in later chapters: Databases: Chapters 3 and 4 BLAST: Chapter 7 Multiple-sequence alignments: Chapters 9 and 10