July 2003
Intermediate to advanced
368 pages
13h 44m
English
Content preview from BLAST
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Chapter 9: BLAST Protocols
WU-BLAST parameters
blastn <genome> <dna> M=1 N=-1 Q=3 R=2 W=9 wordmask=seg lcmask
Expected results
Be on the lookout for repetitive and low-complexity sequence and pseudogenes.
Cross-species alignments are difficult to interpret because many factors impact DNA
evolution. Not all sequences evolve at the same rate, and it is very easy to confuse
signal and noise. It’s a good idea to approach your findings with skepticism.
Sequences that are nearly identical may indicate a very important biological signal, or
they may represent sequencing contamination.
Low-scoring alignments may be coincidental similarities of no biological signifi-
cance. If your search space is large, even high-scoring alignments are expected by
chance. Work out the Karlin-Altschul expectation and search fabricated sequences to
appreciate how frequently false positive alignments occur. That said, some biologi-
cal signals are short and may be buried in the stochastic noise. The best way to deal
with them is to reduce your search space. For example, if you are interested in deter-
mining if there is a short region of interest within the intron of a gene, try aligning
the intron with the orthologous intron from another genome rather than the entire
genome.
If you want to identify orthologs ...
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