- 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.
75
Chapter 5
CHAPTER 5
BLAST
Previous chapters explored what biological sequences are, how they are aligned, and
how similarity is measured. This chapter discusses BLAST itself. What is BLAST?
The simple answer is that it is a set of programs that search sequence databases for
statistically significant similarities. The details of how BLAST searches for similari-
ties aren’t so easily answered. Searching requires multiple steps and many control-
ling parameters. Understanding the theoretical framework will help you design and
interpret BLAST experiments, and give you a foundation for troubleshooting when
your search produces unexpected results.
The Five BLAST Programs
The five traditional BLAST programs are: BLASTN, BLASTP, BLASTX, TBLASTN,
and TBLASTX. BLASTN compares nucleotide sequences to one another (hence the
N). All other programs compare protein sequences (see Table 5-1).
Table 5-1. Traditional BLAST programs
Program Database Query Typical uses
BLASTN Nucleotide Nucleotide Mapping oligonucleotides, cDNAs, and PCR products to a
genome; screening repetitive elements; cross-species
sequence exploration; annotating genomic DNA; clustering
sequencing reads; vector clipping
BLASTP Protein Protein Identifying common regions between proteins; collecting
related proteins for phylogenetic analyses
BLASTX Protein Nucleotide trans-
lated into protein
Finding protein-coding genesin genomicDNA; determining
if a cDNA corresponds to a known protein
TBLASTN Nucleotide trans-
lated into protein
Protein Identifying transcripts, potentially from multiple organ-
isms, similar to a given protein; mapping a protein to
genomic DNA
TBLASTX Nucleotide trans-
lated into protein
Nucleotide trans-
lated into protein
Cross-species gene prediction at the genome or transcript
level; searching for genes missed by traditional methods or
not yet in protein databases
-
No Comment
..................Content has been hidden....................
You can't read the all page of ebook, please click here login for view all page.