- Table of Contents (1/2)
- Table of Contents (2/2)
- Foreword
- Preface
- Part I
- Part II
- Biological Sequences
- Sequence Alignment
- Sequence Similarity
- Part III
- BLAST
- Anatomy of a BLAST Report
- A BLAST Statistics Tutorial
- Basic BLAST Statistics
- Actual Versus Effective Lengths
- The Raw Score and Bit Score
- The Expect of an HSP
- The WU-BLAST P-Value
- Sum Statistics
- An Expect(n) Means That Sum Statistics Were Applied
- Sum Statistics Are Pair-Wise in Their Focus
- The Sum Score
- Effective Length of a BLASTX Query
- Calculating a Sum Score
- Calculating the Pair-Wise Sum P-Value
- Correcting for Multiple Tests
- Correcting for Database Size
- Frame- and Size-Corrected Expects
- Using Statistics to Understand BLAST Results
- Where Did My Oligo Go?
- Basic BLAST Statistics
- 20 Tips to Improve YourBLASTSearches
- 8.1 Don’t Use the Default Parameters
- 8.2 Treat BLAST Searches as Scientific Experiments
- 8.3 Perform Controls, Especially in theTwilightZone
- 8.4 View BLAST Reports Graphically
- 8.5 Use the Karlin-Altschul Equation toDesignExperiments
- 8.6 When Troubleshooting, Read the Footer First
- 8.7 Know When to Use Complexity Filters
- 8.8 Mask Repeats in Genomic DNA
- 8.9 Segment Large Genomic Sequences
- 8.10 Be Skeptical of Hypothetical Proteins
- 8.11 Expect Contaminants in EST Databases
- 8.12 Use Caution When Searching Raw Sequencing Reads
- 8.13 Look for Stop Codons and Frame-Shifts to find Pseudo-Genes
- 8.14 Consider Using Ungapped Alignment for BLASTX, TBLASTN, and TBLASTX
- 8.15 Look for Gaps in Coverage as a Sign ofMissedExons
- 8.16 Parse BLAST Reports with Bioperl
- 8.17 Perform Pilot Experiments
- 8.18 Examine Statistical Outliers
- 8.19 Use links and topcomboN to Make Sense of Alignment Groups
- 8.20 How to Lie with BLAST Statistics
- BLAST Protocols
- BLASTN Protocols (1/3)
- BLASTN Protocols (2/3)
- BLASTN Protocols (3/3)
- BLASTP Protocols
- BLASTX Protocols
- TBLASTN Protocols
- TBLASTX Protocols
- Preventing Stop Codons
- Finding Undocumented Genes in Genomic DNA
- Transcript-Transcript TBLASTX
- Approach
- NCBI-BLAST
- WU-BLAST
- Expected results
- Optimizations and variations
- Part IV
- Installation and Command-Line Tutorial
- BLAST Databases
- Hardware and Software Optimizations
- Distributed Resource Management
- Optimized NCBI-BLAST
- Part V
- NCBI-BLAST Reference
- Usage Statements
- Command-Line Syntax
- blastall Parameters (1/2)
- blastall Parameters (2/2)
- -a [integer]
- -A [integer]
- -b [integer]
- -B [integer]
- -d [database]
- -D [1..23]
- -e [real number]
- -E [integer]
- -f [integer]
- -F [T/F], -F [string]
- -g [T/F]
- -G [integer]
- -i [input file]
- -I [T/F]
- -J [T/F]
- -K [integer]
- -l [file]
- -L [string]
- -m [0..11]
- -M [matrix file]
- -n [T/F]
- -o [output file]
- -p [program name]
- -P [0/1]
- -q [negative integer]
- -Q [1..23]
- -r [integer]
- -R [checkpoint file]
- -S [1..3]
- -t [integer]
- -T [T/F]
- -v [integer]
- -w [integer]
- -W [integer]
- -X [integer]
- -y [integer]
- -Y [real number]
- -z [real number]
- -Z [integer]
- formatdb Parameters
- fastacmd Parameters
- megablast Parameters (1/2)
- megablast Parameters (2/2)
- -a [integer]
- -A [integer]
- -b [integer]
- -d [string]
- -D [0..3]
- -e [real number]
- -E [integer]
- -f [T/F]
- -F [T/F] [string]
- -G [integer]
- -H [integer]
- -i [file]
- -I [T/F]
- -l [file]
- -L [string]
- -m [0..11]
- -M [integer]
- -n [T/F]
- -N [0,1,2]
- -o [file]
- -p [real number]
- -P [integer]
- -q [negative integer]
- -Q [file]
- -r [integer]
- -R [T/F]
- -s [integer]
- -S [0..3]
- -t [16,18,21]
- -T [T/F]
- -U [T/F]
- -v [integer]
- -W [integer]
- -X [integer]
- -y [integer]
- -z [real number]
- -Z [integer]
- bl2seq Parameters
- -a [file]
- -A [T/F]
- -d [real number]
- -D [0/1]
- -e [real number]
- -E [integer]
- -F [T/F] [string]
- -g [T/F]
- -G [integer]
- -i [file]
- -I [integer],[integer]
- -j [file]
- -J [integer],[integer]
- -m [T/F]
- -M [string]
- -o [file]
- -p [string]
- -q [negative integer]
- -r [integer]
- -S [1..3]
- -t [integer]
- -T [T/F]
- -U [T/F]
- -W [integer]
- -X [integer]
- -Y [real number]
- blastpgp Parameters (PSI-BLAST andPHIBLAST) (1/2)
- blastpgp Parameters (PSI-BLAST andPHIBLAST) (2/2)
- PSI-BLAST
- PHI-BLAST
- -a [integer]
- -A [integer]
- -b [integer]
- -B [file]
- -c [integer]
- -C [file]
- -d [string]
- -e [real]
- -E [integer]
- -f [integer]
- -F [string]
- -g [T/F]
- -G [integer]
- -h [real number]
- -H [integer]
- -i [file]
- -I [T/F]
- -j [integer]
- -J [T/F]
- -k [file]
- -K [integer]
- -l [string]
- -L [integer]
- -m [0..9]
- -M [string]
- -N [real number]
- -o [file]
- -O [file]
- -p [string]
- -Q [file]
- -R [file]
- -s [T/F]
- -S [integer]
- -t [T/F]
- -T [T/F]
- -U [T/F]
- -v [integer]
- -W [1..3]
- -X [integer]
- -y [real number]
- -Y [real number]
- -z [real number]
- -Z [integer]
- blastclust Parameters
- WU-BLAST Reference
- Usage Statements
- Command-Line Syntax
- WU-BLAST Parameters (1/3)
- WU-BLAST Parameters (2/3)
- WU-BLAST Parameters (3/3)
- altscore=[string]
- B=[integer]
- bottom
- cpus=[integer]
- dbrecmax=[integer]
- dbrecmin=[integer]
- E=[number]
- E2=[number]
- echofilter
- errors
- filter=[string]
- gapE2=[number]
- gapH=[number]
- gapK=[number]
- gapL=[number]
- gapS2=[integer]
- gapsepqmax=[int]
- gapsepsmax=[int]
- gapX
- gi
- golf=[number]
- golmax=[integer]
- gspmax=[integer]
- H=[number]
- hspmax=[integer]
- hitdist=[integer]
- hspsepqmax=[int]
- hspsepsmax=[int]
- K=[number]
- kap
- L=[number]
- lcfilter
- lcmask
- links
- M=[integer]
- maskextra=[integer]
- matrix=[file]
- N=[integer]
- nogap
- nonnegok
- nosegs
- notes
- novalidctxok
- nwlen=[integer]
- nwstart=[integer]
- o=[file]
- olf=[number]
- olmax=[integer]
- postsw
- Q=[integer]
- qoffset=[integer]
- qrecmax=[integer]
- Qrecmin=[integer]
- R=[integer]
- restest
- S=[integer]
- mS2=[integer]
- seqtest
- span, span1, span2
- T=[integer]
- top
- topcomboN=[integer]
- V=[integer]
- warnings
- wink=[integer]
- wordmask=[method]
- W=[integer]
- X=[integer]
- Y=[number]
- Z=[number]
- xdformat Parameters
- xdget Parameters
- NCBI-BLAST Reference
- Part VI
- NCBI Display Formats
- Brief Descriptions
- Detailed Descriptions and Examples
- Option 0: Pairwise Alignments
- Query-Anchored Alignments
- Option 1: Query-Anchored Showing Identities
- Option 2: Query-Anchored, No Identities
- Option 3: Flat Query-Anchored Showing Identities
- Option 4: Flat Query-Anchored, No Identities
- Option 5: Query-Anchored, No Identities, and Blunt Ends
- Option 6: Flat Query-Anchored, No Identities, and Blunt Ends
- Option 7: XML
- Option 8: Tabular, Without Comment Lines
- Option 9: Tabular, with Comment Lines
- Option 10: ASN.1 Text Format
- Option 11: ASN.1 Binary Format
- Nucleotide Scoring Schemes
- NCBI-BLAST Scoring Schemes
- blast-imager.pl
- blast2table.pl
- NCBI Display Formats
- Glossary (1/2)
- Glossary (2/2)
- Index (1/5)
- Index (2/5)
- Index (3/5)
- Index (4/5)
- Index (5/5)
This is the Title of the Book, eMatter Edition
Copyright © 2012 O’Reilly & Associates, Inc. All rights reserved.
Further Reading
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87
but a warning is issued that informs you that you can provide your own values for
lambda, k, and H on the command line.
NCBI’s version of BLASTN doesn’t contain gapped values for lambda; lambda is
always calculated directly from the match/mismatch scores. Because of this, equiva-
lent alignments may have much higher bit scores (and lower E-values) in NCBI-
BLAST than WU-BLAST, even if their match/mismatch scores are identical.
Further Reading
The following papers describe BLAST algorithms in more detail:
Altschul, Stephen F., Warren Gish, Webb Miller, Eugene W. Myers, and David J.
Lipman (1990). “Basic local alignment search tool,” Journal of Molecular Biol-
ogy, 215:403-10.
Altschul, Stephen F., Thomas L. Madden, Alejandro A. Schaffer, Jinghui Zhang,
Zheng Zhang, Webb Miller, and David J. Lipman (1997), “Gapped BLAST and
PSI-BLAST: a new generation of protein database search programs,” Nucleic
Acids Research, 25:3389-3402.
Gish, Warren and David J. States (1993). “Identification of protein coding regions by
database similarity search,” Nature Genet., 3:266-72.
For the latest information on NCBI-BLAST and other helpful documents, see the
official web site at http://ncbi.nlm.nih.gov/BLAST.
The WU-BLAST web site (http://blast.wustl.edu) has several useful documents on
BLAST in general, and includes pages specifically for WU-BLAST users. One of its
most interesting documents is a thorough manual for the classic, ungapped version
of BLAST 1.4 (http://blast.wustl.edu/doc/blast1.pdf).
-
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