BLAST (Basic Local Alignment Search Tool)
Introduction
The Basic Local Alignment Search Tool (BLAST) is a widely used bioinformatics program designed for comparing primary biological sequence information. This includes nucleotide sequences of DNA and RNA, as well as protein sequences. BLAST is instrumental in identifying homologous sequences and inferring functional and evolutionary relationships between sequences. It is a fundamental tool in the field of computational biology and is extensively used in genomics, proteomics, and evolutionary biology.
History and Development
BLAST was developed in the late 1980s by Stephen Altschul, Warren Gish, Webb Miller, Eugene Myers, and David J. Lipman at the National Center for Biotechnology Information (NCBI). The initial publication in 1990 introduced a novel approach to sequence alignment that was significantly faster than previous methods, such as the Smith-Waterman algorithm. The development of BLAST marked a pivotal moment in bioinformatics, allowing researchers to perform sequence alignments on a much larger scale.
Algorithm and Methodology
BLAST operates by dividing the query sequence into smaller "words" and searching for these words in a database of sequences. The algorithm uses a heuristic approach to identify regions of local similarity, which are then extended to generate alignments. This method is computationally efficient, making it possible to search large databases quickly.
Word Size
The word size is a critical parameter in BLAST, influencing both the sensitivity and speed of the search. A smaller word size increases sensitivity but requires more computational resources, while a larger word size speeds up the process but may miss some alignments. The default word size varies depending on the type of sequence being analyzed (e.g., nucleotide or protein).
Scoring Matrices
BLAST uses scoring matrices, such as PAM or BLOSUM, to evaluate the quality of alignments. These matrices assign scores to matches, mismatches, and gaps, reflecting the likelihood of evolutionary substitutions. The choice of matrix can significantly affect the results, and selecting an appropriate matrix is crucial for accurate sequence alignment.
E-value
The Expectation value, or E-value, is a statistical measure used in BLAST to assess the significance of an alignment. It represents the number of alignments with a score equal to or better than the observed score that would be expected to occur by chance. A lower E-value indicates a more significant alignment.
Types of BLAST Programs
BLAST offers several variations tailored to specific types of sequence comparisons:
BLASTN
BLASTN is used for nucleotide-to-nucleotide sequence comparisons. It is commonly employed in genomics to identify homologous DNA sequences.
BLASTP
BLASTP performs protein-to-protein sequence comparisons. It is widely used in proteomics to find similar protein sequences and infer functional relationships.
BLASTX
BLASTX translates a nucleotide query sequence into all possible protein sequences and compares them against a protein database. This is useful for identifying potential protein-coding regions in nucleotide sequences.
TBLASTN
TBLASTN compares a protein query sequence against a nucleotide database translated in all six reading frames. It is often used to identify homologous sequences in genomic DNA.
TBLASTX
TBLASTX translates both the query and database nucleotide sequences into proteins and performs a protein-to-protein comparison. This approach is helpful for identifying conserved regions in non-coding DNA.
Applications
BLAST has a wide range of applications in various fields of biological research:
Genomics
In genomics, BLAST is used to annotate genomes by identifying genes and predicting their functions. It facilitates the comparison of newly sequenced genomes with existing databases, aiding in the discovery of novel genes.
Proteomics
In proteomics, BLAST helps in the identification and characterization of proteins. It is used to predict protein functions, identify protein families, and study protein evolution.
Evolutionary Biology
BLAST is a valuable tool in evolutionary biology for studying phylogenetic relationships. By comparing sequences from different organisms, researchers can infer evolutionary lineages and trace the history of genes.
Medical Research
In medical research, BLAST is used to identify genetic mutations associated with diseases. It aids in the development of diagnostic tools and the discovery of potential therapeutic targets.
Limitations and Challenges
Despite its widespread use, BLAST has certain limitations. The heuristic nature of the algorithm means that it may miss some alignments, particularly those with low similarity. Additionally, the choice of parameters, such as word size and scoring matrix, can significantly impact the results. Researchers must carefully select these parameters to ensure accurate and meaningful alignments.
Future Developments
Ongoing research in bioinformatics aims to enhance the capabilities of BLAST and develop new algorithms for sequence alignment. Advances in computational power and machine learning are expected to improve the sensitivity and speed of sequence searches, enabling more comprehensive analyses of biological data.