1. Introduction

   1. Chapter 1 Hello BLAST

      1. What Is BLAST?
      2. Using NCBI-BLAST
      3. Alternate Output Formats
      4. Alternate Alignment Views
      5. The Next Step
      6. Further Reading

2. Theory

   1. Chapter 2 Biological Sequences

      1. The Central Dogma of Molecular Biology
      2. Evolution
      3. Genomes and Genes
      4. Biological Sequences and Similarity
      5. Further Reading

   2. Chapter 3 Sequence Alignment

      1. Global Alignment: Needleman-Wunsch
      2. Local Alignment: Smith-Waterman
      3. Dynamic Programming
      4. Algorithmic Complexity
      5. Global Versus Local
      6. Variations
      7. Final Thoughts
      8. Further Reading

   3. Chapter 4 Sequence Similarity

      1. Introduction to Information Theory
      2. Amino Acid Similarity
      3. Scoring Matrices
      4. Target Frequencies, lambda, and H
      5. Sequence Similarity
      6. Karlin-Altschul Statistics
      7. Sum Statistics and Sum Scores
      8. Further Reading

3. Practice

   1. Chapter 5 BLAST

      1. The Five BLAST Programs
      2. The BLAST Algorithm
      3. Further Reading

   2. Chapter 6 Anatomy of a BLAST Report

      1. Basic Structure
      2. Alignments

   3. Chapter 7 A BLAST Statistics Tutorial

      1. Basic BLAST Statistics
      2. Using Statistics to Understand BLAST Results
      3. Where Did My Oligo Go?

   4. Chapter 8 20 Tips to Improve Your BLAST Searches

      1. Don't Use the Default Parameters
      2. Treat BLAST Searches as Scientific Experiments
      3. Perform Controls, Especially in the Twilight Zone
      4. View BLAST Reports Graphically
      5. Use the Karlin-Altschul Equation to Design Experiments
      6. When Troubleshooting, Read the Footer First
      7. Know When to Use Complexity Filters
      8. Mask Repeats in Genomic DNA
      9. Segment Large Genomic Sequences
      10. Be Skeptical of Hypothetical Proteins
      11. Expect Contaminants in EST Databases
      12. Use Caution When Searching Raw Sequencing Reads
      13. Look for Stop Codons and Frame-Shifts to find Pseudo-Genes
      14. Consider Using Ungapped Alignment for BLASTX, TBLASTN, and TBLASTX
      15. Look for Gaps in Coverage as a Sign of Missed Exons
      16. Parse BLAST Reports with Bioperl
      17. Perform Pilot Experiments
      18. Examine Statistical Outliers
      19. Use links and topcomboN to Make Sense of Alignment Groups
      20. How to Lie with BLAST Statistics

   5. Chapter 9 BLAST Protocols

      1. BLASTN Protocols
      2. BLASTP Protocols
      3. BLASTX Protocols
      4. TBLASTN Protocols
      5. TBLASTX Protocols

4. Industrial-Strength BLAST

   1. Chapter 10 Installation and Command-Line Tutorial

      1. NCBI-BLAST Installation
      2. WU-BLAST Installation
      3. Command-Line Tutorial
      4. Editing Scoring Matrices

   2. Chapter 11 BLAST Databases

      1. FASTA Files
      2. BLAST Databases
      3. Sequence Databases
      4. Sequence Database Management Strategies

   3. Chapter 12 Hardware and Software Optimizations

      1. The Persistence of Memory
      2. CPUs and Computer Architecture
      3. Compute Clusters
      4. Distributed Resource Management
      5. Software Tricks
      6. Optimized NCBI-BLAST

5. BLAST Reference

   1. Chapter 13 NCBI-BLAST Reference

      1. Usage Statements
      2. Command-Line Syntax
      3. blastall Parameters
      4. formatdb Parameters
      5. fastacmd Parameters
      6. megablast Parameters
      7. bl2seq Parameters
      8. blastpgp Parameters (PSI-BLAST and PHI-BLAST)
      9. blastclust Parameters

   2. Chapter 14 WU-BLAST Reference

      1. Usage Statements
      2. Command-Line Syntax
      3. WU-BLAST Parameters
      4. xdformat Parameters
      5. xdget Parameters

6. Appendixes

   1. Appendix A NCBI Display Formats

      1. Brief Descriptions
      2. Detailed Descriptions and Examples

   2. Appendix B Nucleotide Scoring Schemes

   3. Appendix C NCBI-BLAST Scoring Schemes

      1. NCBI-BLAST Matrices and Gap Costs

   4. Appendix D blast-imager.pl

   5. Appendix E blast2table.pl

1. Glossary

2. Colophon

Our look is the result of reader comments, our own experimentation, and feedback from distribution channels. Distinctive covers complement our distinctive approach to technical topics, breathing personality and life into potentially dry subjects. The animal on the cover of BLAST is a coelacanth.The modern coelacanth, Latimeria chalumnae, is the sole living representative of Actinista, an ancient family of lobe-finned fish. The first live coelacanth was discovered off the east coast of South Africa in 1938 and caused a stir in the scientific community because they were believed to have been extinct for approximately 80 million years. A second, closely related species of coelacanths was found off the coast of Sulawesi in 1998.

Coelacanths grow to about 6 feet long; weigh up to 150 pounds; and are covered with bony, dark blue scales flecked with white. Coelacanths are unlike other living fish in that they have a three-lobed tail, fleshy fins, and a partially developed vertebral column. They are the only living vertebrate with a functional intercranial joint. It's this joint that allows the coelacanth's jaws to open exceptionally wide when inhaling its prey. Coelacanths are opportunistic predators (they eat whatever they can find) and are ovoviviparous (they give birth to live young).

From an evolutionary perspective, coelacanths are unique, and they provide a key perspective on the evolution of many different genes in vertebrate species. With their front and rear paired fins, which move in a similar fashion to the arms and legs of land vertebrates, coelacanths may be one of the closest links to those vertebrates that first crawled out of the sea to live on land more than 350 million years ago. Mary Anne Weeks Mayo was the production editor and proofreader, and Ann Schirmer was the copyeditor for BLAST. Derek DiMatteo, and Darren Kelly provided quality control. Lucie Haskins wrote the index.

Emma Colby designed the cover of this book, based on a series design by Edie Freedman. The cover image is an original illustration created by Lorrie LeJeune. Emma produced the cover layout with QuarkXPress 4.1 using Adobe's ITC Garamond font.

David Futato designed the interior layout. This book was converted by Julie Hawks to FrameMaker 5.5.6 with a format conversion tool created by Erik Ray, Jason McIntosh, Neil Walls, and Mike Sierra that uses Perl and XML technologies. The text font is Linotype Birka; the heading font is Adobe Myriad Condensed; and the code font is LucasFont's TheSans Mono Condensed. The illustrations that appear in the book were produced by Robert Romano and Jessamyn Read using Macromedia FreeHand 9 and Adobe Photoshop 6. This colophon was compiled by Lorrie LeJeune.