Finding Homologs in Amino Acid Sequences Using Network BLAST Searches

Abstract

BLAST, the Basic Local Alignment Search Tool, is used more frequently than any other biosequence database search program. We show how to run searches on the Web, and demonstrate how to increase performance by fine-tuning arguments for a specific research project. We offer guidance for interpreting results, statistical significance and biological relevance issues, and suggest complementary analyses. This unit covers both protein-to-protein (blastp) searches and translated searches (blastx, tblastn, tfastx). blastx conceptually translates the query sequence and tblastn translates all nucleotide sequences in a database, while tblastx translates both the query and the database sequences into amino acid sequences. © 2017 by John Wiley & Sons, Inc.

Installing, Maintaining, and Using a Local Copy of BLAST for Compute Cluster or Workstation Use

“…The utility of BLAST search results critically depends on the selection of the databases to searched and whether they are up to date. For the selection of databases to serve your specific objectives, refer to Ladunga, (,b) and Pearson (2013b), and a recent publication about the NCBI services and databases (NCBI Resource Coordinators, ). Large organizations are expected to perform fresh downloads of the major databases from NCBI every night.…”

“…BLAST, the Basic Local Alignment Search Tool (Altschul et al., ; Altschul, Gish, Miller, Myers, & Lipman, ; Ladunga, ,b), is the first and most widely used computational tool in the characterization of nucleic acid and amino acid sequences. BLAST can predict the function of novel sequences, identify phylogenetic relationships, assist in the prediction of three‐dimensional structures of proteins, locate conserved regions, and aid in the design of targeted genome modifications or RNA interference, among many other possible uses.…”

Finding Protein and Nucleotide Similarities with FASTA

“…There are three widely used sets of programs for searching protein and DNA sequences, the BLAST package [ UNIT here (Ladunga, ) and UNIT here (Ladunga, )], HMMER (Eddy, ; Finn et al., ), and the FASTA package. These programs do similar things—they compare a query sequence to a library of sequences, calculating tens of thousands to millions of similarity scores, and report back the library sequences that are most similar to the query.…”

“…The FASTA package provides programs for searching protein, DNA, or translated DNA sequence databases using proteins, DNA, translated DNA, or short peptides as queries. Table summarizes the programs that should be used for various analysis problems, and corresponding programs, if any, in the BLAST package [ UNIT here (Ladunga, ) and UNIT here (Ladunga, )]. In general, if one has a protein sequence, one should use fasta36 or ssearch36 [a Smith‐Waterman implementation; Smith and Waterman, ; also see UNIT here (Ropelewski et al., )].…”

“…The FASTA software package provides a comprehensive set of programs (Table ) for protein and DNA sequence comparison. While the FASTA programs are not as fast as the BLAST programs [ UNIT here (Ladunga, ) and UNIT here (Ladunga, )], they can be equally sensitive, and, because they calculate statistical parameters from the distribution of similarity scores calculated during the search, they can provide more accurate statistical estimates using a wide range of scoring parameters. Programs in the FASTA package offer a broad range of speed, sensitivity, and alignment and statistical accuracy for similarity searches and statistical analysis.…”