July 2003
Intermediate to advanced
368 pages
13h 44m
English
Content preview from BLAST
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Chapter 9: BLAST Protocols
The values for X and S2 are calculated based on the default BLOSUM62 matrix. The
most negative score is –4, so
X=10 allows at least two mismatches. The average match
score is about 5, and the average mismatch score is about –1.5 (you can estimate
these scores quickly by looking at common amino acids such as alanine and gly-
cine).
S2=65 corresponds to a 90 percent identity alignment of 15 amino acids.
NCBI-BLAST parameters
blastall -p blastx -d <db> -i <dna> -f 999 -A 4 -G 32767 -E 32767
WU-BLAST parameters
blastx <db> <dna> filter=seg W=6 WINK=6 nogap
WU-BLAST 1.4 parameters
blastx <db> <dna> filter=seg W=5 S2=65 X=10
Expected results
Not all exons will be hit using such parameters, but with this mapping experiment,
the general coordinates are of greatest interest. If you need an accurate alignment,
this search can be followed by a more sensitive search using a serial strategy or
bl2seq.
How much faster are these searches? Speed depends on sequence length and con-
tent, but as a general observation for sequences in the 50-200 Kb range, if the default
NCBI parameter set is considered 1x, the fastest NCBI-BLAST is 6-8x, WU-BLAST is
100-500x, and the classic WU-BLAST 1.4 is 50-150x.
Optimizations and variations
Other alignment algorithms such as BLAT (yes, the
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