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Multiple Sequence Alignment Methods pp 75–101Cite as

BLAST and FASTA Similarity Searching for Multiple Sequence Alignment

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Part of the book series: Methods in Molecular Biology ((MIMB,volume 1079))

Abstract

BLAST, FASTA, and other similarity searching programs seek to identify homologous proteins and DNA sequences based on excess sequence similarity. If two sequences share much more similarity than expected by chance, the simplest explanation for the excess similarity is common ancestry—homology. The most effective similarity searches compare protein sequences, rather than DNA sequences, for sequences that encode proteins, and use expectation values, rather than percent identity, to infer homology. The BLAST and FASTA packages of sequence comparison programs provide programs for comparing protein and DNA sequences to protein databases (the most sensitive searches). Protein and translated-DNA comparisons to protein databases routinely allow evolutionary look back times from 1 to 2 billion years; DNA:DNA searches are 5–10-fold less sensitive. BLAST and FASTA can be run on popular web sites, but can also be downloaded and installed on local computers. With local installation, target databases can be customized for the sequence data being characterized. With today’s very large protein databases, search sensitivity can also be improved by searching smaller comprehensive databases, for example, a complete protein set from an evolutionarily neighboring model organism. By default, BLAST and FASTA use scoring strategies target for distant evolutionary relationships; for comparisons involving short domains or queries, or searches that seek relatively close homologs (e.g. mouse–human), shallower scoring matrices will be more effective. Both BLAST and FASTA provide very accurate statistical estimates, which can be used to reliably identify protein sequences that diverged more than 2 billion years ago.

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Notes

  1. 1.

    The programs listed in Tables 1 and 2 are part of the NCBI BLAST+ distribution [1]. An earlier version of the BLAST distribution used the blastall program with the -p blastp option to specify the specific program.

  2. 2.

    Previous versions of BLAST provided bl2seq to compare two sequences in FASTA format. The BLAST programs now provide the “Blast2Sequences” mode by using -subject option, rather than the -db option.

  3. 3.

    The FASTA programs provide a variable scoring matrix option that shifts the scoring matrix for shorter query sequences. The BLAST programs provide the -task blastp-short or -task blastn-short for short protein:protein and DNA:DNA searches.

  4. 4.

    BLAST’s percent positive counts aligned residues with a score > 0; FASTA’s fraction similar includes aligned residues with scores ≥ 0.

  5. 5.

    The BLAST programs use a slightly different formulation of the Expect value; rather than using the number of entries in the database, BLAST uses the combined length of all the sequences in the database. For average length proteins, the result of the two calculations is identical.

  6. 6.

    There are more than 2.5 million E. coli protein sequences from 200+ genomes available from the NCBI protein databases.

  7. 7.

    Similar results are found with blastp with the PAM70 matrix, though the less stringent gap penalties used by blastp produce longer alignments.

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Authors and Affiliations

  1. Department of Biochemistry and Molecular Genetics, University of Virginia School of Medicine, Charlottesville, VA, USA

    William R. Pearson

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  1. William R. Pearson
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Editors and Affiliations

  1. Dept. of Electrical Engineering, University of Nebraska-Lincoln, Lincoln, Nebraska, USA

    David J Russell

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Pearson, W.R. (2014). BLAST and FASTA Similarity Searching for Multiple Sequence Alignment. In: Russell, D. (eds) Multiple Sequence Alignment Methods. Methods in Molecular Biology, vol 1079. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-646-7_5

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  • DOI: https://doi.org/10.1007/978-1-62703-646-7_5

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  • Publisher Name: Humana Press, Totowa, NJ

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