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    BLAST N FASTA

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    Anshuman Panda
    The document discusses pairwise alignment methods in bioinformatics, specifically global and local alignments, highlighting the Needelman-Wunch and Smith-Waterman algorithms. It introduces heuristic methods like FASTA and BLAST for sequence searching, emphasizing their differences in speed, user-friendliness, and statistical rigor. The document also details the processes involved in both FASTA and BLAST algorithms, including word matching and alignment optimization.

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    BLAST N FASTA

    Uploaded by

    Anshuman Panda
    2 views55 pages
    The document discusses pairwise alignment methods in bioinformatics, specifically global and local alignments, highlighting the Needelman-Wunch and Smith-Waterman algorithms. It introduces heuristic methods like FASTA and BLAST for sequence searching, emphasizing their differences in speed, user-friendliness, and statistical rigor. The document also details the processes involved in both FASTA and BLAST algorithms, including word matching and alignment optimization.

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    The document discusses pairwise alignment methods in bioinformatics, specifically global and local alignments, highlighting the Needelman-Wunch and Smith-Waterman algorithms. It introduces heuristic methods like FASTA and BLAST for sequence searching, emphasizing their differences in speed, user-friendliness, and statistical rigor. The document also details the processes involved in both FASTA and BLAST algorithms, including word matching and alignment optimization.

    Copyright:

    © All Rights Reserved
    Download as PPT, PDF, TXT or read online from Scribd
    Download as ppt, pdf, or txt
    2 views55 pages

    BLAST N FASTA

    Uploaded by

    Anshuman Panda
    The document discusses pairwise alignment methods in bioinformatics, specifically global and local alignments, highlighting the Needelman-Wunch and Smith-Waterman algorithms. It introduces heuristic methods like FASTA and BLAST for sequence searching, emphasizing their differences in speed, user-friendliness, and statistical rigor. The document also details the processes involved in both FASTA and BLAST algorithms, including word matching and alignment optimization.

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    © All Rights Reserved
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    You are on page 1of 55

    BLAST and FASTA

    1
    Pairwise Alignment

    Global Local
    • Best score from among • Best score from among
    alignments of full-length alignments of partial
    sequences sequences
    • Needelman-Wunch • Smith-Waterman
    algorithm algorithm

    8
    Local vs. Global Alignment
    • Global Alignment
    --T—-CC-C-AGT—-TATGT-CAGGGGACACG—A-GCATGCAGA-GAC
    | || | || | | | ||| || | | | | |||| |
    AATTGCCGCC-GTCGT-T-TTCAG----CA-GTTATG—T-CAGAT--C

    • Local Alignment—better
    alignment to find conserved
    segment tccCAGTTATGTCAGgggacacgagcatgcagagac
    ||||||||||||
    aattgccgccgtcgttttcagCAGTTATGTCAGatc

    9
    Why do we need local alignments?

    • To compare a short sequence to a large one.

    • To compare a single sequence to an entire


    database

    • To compare a partial sequence to the whole.

    10
    11
    Heuristic Methods: FASTA and BLAST

    FASTA
    • First fast sequence searching algorithm for
    comparing a query sequence against a database.

    BLAST
    • Basic Local Alignment Search Technique
    improvement of FASTA: Search speed, ease of
    use, statistical rigor.
    12
    FASTA and BLAST
    • Basic idea: a good alignment contains
    subsequences of absolute identity (short lengths
    of exact matches):

    – First, identify very short exact matches.


    – Next, the best short hits from the first step are
    extended to longer regions of similarity.
    – Finally, the best hits are optimized.

    13
    FASTA
    Derived from logic of the dot plot
    – compute best diagonals from all frames of
    alignment
    The method looks for exact matches between
    words in query and test sequence
    – DNA words are usually 6 nucleotides long
    – protein words are 2 amino acids long

    14
    FASTA Algorithm

    17
    Makes Longest Diagonal
    After all diagonals are found, tries to join
    diagonals by adding gaps

    Computes alignments in regions of best


    diagonals

    18
    FASTA Alignments

    19
    FASTA Results - Histogram
    !!SEQUENCE_LIST 1.0
    (Nucleotide) FASTA of: b2.seq from: 1 to: 693 December 9, 2002 14:02
    TO: /u/browns02/Victor/Search-set/*.seq Sequences: 2,050 Symbols:
    913,285 Word Size: 6
    Searching with both strands of the query.
    Scoring matrix: GenRunData:fastadna.cmp
    Constant pamfactor used
    Gap creation penalty: 16 Gap extension penalty: 4

    Histogram Key:
    Each histogram symbol represents 4 search set sequences
    Each inset symbol represents 1 search set sequences
    z-scores computed from opt scores
    z-score obs exp
    (=) (*)
    < 20 0 0:
    22 0 0:
    24 3 0:=
    26 2 0:=
    28 5 0:==
    30 11 3:*==
    32 19 11:==*==
    34 38 30:=======*==
    36 58 61:===============*
    38 79 100:==================== *
    40 134 140:==================================*
    42 167 171:==========================================*
    44 205 189:===============================================*==== 20
    46 209 192:===============================================*=====
    48 177 184:=============================================*
    FASTA Results - List
    The best scores are: init1 initn opt z-sc E(1018780)..

    SW:PPI1_HUMAN Begin: 1 End: 269


    ! Q00169 homo sapiens (human). phosph... 1854 1854 1854 2249.3 1.8e-117
    SW:PPI1_RABIT Begin: 1 End: 269
    ! P48738 oryctolagus cuniculus (rabbi... 1840 1840 1840 2232.4 1.6e-116
    SW:PPI1_RAT Begin: 1 End: 270
    ! P16446 rattus norvegicus (rat). pho... 1543 1543 1837 2228.7 2.5e-116
    SW:PPI1_MOUSE Begin: 1 End: 270
    ! P53810 mus musculus (mouse). phosph... 1542 1542 1836 2227.5 2.9e-116
    SW:PPI2_HUMAN Begin: 1 End: 270
    ! P48739 homo sapiens (human). phosph... 1533 1533 1533 1861.0 7.7e-96
    SPTREMBL_NEW:BAC25830 Begin: 1 End: 270
    ! Bac25830 mus musculus (mouse). 10, ... 1488 1488 1522 1847.6 4.2e-95
    SP_TREMBL:Q8N5W1 Begin: 1 End: 268
    ! Q8n5w1 homo sapiens (human). simila... 1477 1477 1522 1847.6 4.3e-95
    SW:PPI2_RAT Begin: 1 End: 269
    ! P53812 rattus norvegicus (rat). pho... 1482 1482 1516 1840.4 1.1e-94

    21
    FASTA Results - Alignment
    SCORES Init1: 1515 Initn: 1565 Opt: 1687 z-score: 1158.1 E(): 2.3e-58
    >>GB_IN3:DMU09374 (2038 nt)
    initn: 1565 init1: 1515 opt: 1687 Z-score: 1158.1 expect(): 2.3e-58
    66.2% identity in 875 nt overlap
    (83-957:151-1022)

    60 70 80 90 100 110
    u39412.gb_pr CCCTTTGTGGCCGCCATGGACAATTCCGGGAAGGAAGCGGAGGCGATGGCGCTGTTGGCC
    || ||| | ||||| | ||| |||||
    DMU09374 AGGCGGACATAAATCCTCGACATGGGTGACAACGAACAGAAGGCGCTCCAACTGATGGCC
    130 140 150 160 170 180

    120 130 140 150 160 170


    u39412.gb_pr GAGGCGGAGCGCAAAGTGAAGAACTCGCAGTCCTTCTTCTCTGGCCTCTTTGGAGGCTCA
    ||||||||| || ||| | | || ||| | || || ||||| ||
    DMU09374 GAGGCGGAGAAGAAGTTGACCCAGCAGAAGGGCTTTCTGGGATCGCTGTTCGGAGGGTCC
    190 200 210 220 230 240

    180 190 200 210 220 230


    u39412.gb_pr TCCAAAATAGAGGAAGCATGCGAAATCTACGCCAGAGCAGCAAACATGTTCAAAATGGCC
    ||| | ||||| || ||| |||| | || | |||||||| || ||| ||
    DMU09374 AACAAGGTGGAGGACGCCATCGAGTGCTACCAGCGGGCGGGCAACATGTTTAAGATGTCC
    250 260 270 280 290 300

    240 250 260 270 280 290


    u39412.gb_pr AAAAACTGGAGTGCTGCTGGAAACGCGTTCTGCCAGGCTGCACAGCTGCACCTGCAGCTC
    |||||||||| ||||| | |||||| |||| ||| || ||| || |
    DMU09374 AAAAACTGGACAAAGGCTGGGGAGTGCTTCTGCGAGGCGGCAACTCTACACGCGCGGGCT 22
    310 320 330 340 350 360
    FASTA Format
    • simple format used by almost all programs
    • [>] header line with a [hard return] at end
    • Sequence (no specific requirements for line
    length, characters, etc)

    >URO1 uro1.seq Length: 2018 November 9, 2000 11:50 Type: N Check: 3854 ..
    CGCAGAAAGAGGAGGCGCTTGCCTTCAGCTTGTGGGAAATCCCGAAGATGGCCAAAGACA
    ACTCAACTGTTCGTTGCTTCCAGGGCCTGCTGATTTTTGGAAATGTGATTATTGGTTGTT
    GCGGCATTGCCCTGACTGCGGAGTGCATCTTCTTTGTATCTGACCAACACAGCCTCTACC
    CACTGCTTGAAGCCACCGACAACGATGACATCTATGGGGCTGCCTGGATCGGCATATTTG
    TGGGCATCTGCCTCTTCTGCCTGTCTGTTCTAGGCATTGTAGGCATCATGAAGTCCAGCA
    GGAAAATTCTTCTGGCGTATTTCATTCTGATGTTTATAGTATATGCCTTTGAAGTGGCAT
    CTTGTATCACAGCAGCAACACAACAAGACTTTTTCACACCCAACCTCTTCCTGAAGCAGA
    TGCTAGAGAGGTACCAAAACAACAGCCCTCCAAACAATGATGACCAGTGGAAAAACAATG
    GAGTCACCAAAACCTGGGACAGGCTCATGCTCCAGGACAATTGCTGTGGCGTAAATGGTC
    CATCAGACTGGCAAAAATACACATCTGCCTTCCGGACTGAGAATAATGATGCTGACTATC 23
    CCTGGCCTCGTCAATGCTGTGTTATGAACAATCTTAAAGAACCTCTCAACCTGGAGGCTT
    BLAST
    • Basic Local Alignment Search Tool
    – Altschul et al. 1990,1994,1997
    • Heuristic method for local alignment
    • Designed specifically for database searches
    • Based on the same assumption as FASTA
    that good alignments contain short lengths
    of exact matches
    24
    BLAST
    • Both BLAST and FASTA search for local
    sequence similarity - indeed they have exactly
    the same goals, though they use somewhat
    different algorithms and statistical approaches.

    • BLAST benefits
    – Speed
    – User friendly
    – Statistical rigor
    – More sensitive
    25
    BLAST Searches GenBank
    [BLAST= Basic Local Alignment Search Tool]
    The NCBI BLAST web server lets you compare your
    query sequence to various sections of GenBank:
    – nr = non-redundant (main sections)
    – month = new sequences from the past few weeks
    – refseq_rna
    – RNA entries from NCBI's Reference Sequence project
    – refseq_genomic
    – Genomic entries from NCBI's Reference Sequence project
    – ESTs
    – Taxon = e.g., human, Drososphila, yeast, E. coli
    – proteins (by automatic translation)
    – pdb = Sequences derived from the 3-dimensional structure
    from Brookhaven Protein Data Bank
    26
    BLAST
    • Uses word matching like FASTA
    • Similarity matching of words (3 amino acids, 11
    bases)
    – does not require identical words.
    • If no words are similar, then no alignment
    – Will not find matches for very short sequences

    • Does not handle gaps well


    • “gapped BLAST” is somewhat better
    27
    BLAST Algorithm

    28
    BLAST Word Matching

    MEAAVKEEISVEDEAVDKNI
    MEA
    EAA Break query
    AAV
    AVK into words:
    VKE
    KEE
    EEI
    EIS
    ISV Break database
    ...
    sequences
    into words:

    29
    Find locations of matching words
    in database sequences

    ELEPRRPRYRVPDVLVADPPIARLSVSGRDENSVELTMEAT
    MEA
    EAA TDVRWMSETGIIDVFLLLGPSISDVFRQYASLTGTQALPPLFSLGYHQSRWNY
    AAV IWLDIEEIHADGKRYFTWDPSRFPQPRTMLERLASKRRVKLVAIVDPH
    AVK
    KLV
    KEE
    EEI
    EIS
    ISV

    30
    Extend hits one base at a time

    31
    BLAST variants

    32
    33
    34
    35
    Understanding BLAST output

    36
    37
    38
    39
    40
    41
    44
    45

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