Fr 410: Introduction History, Database Biology, Sequence Formats

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• Fr 4/10 Introduction History, Database
  Biology, Sequence Formats
• Fr 11/10 Pairwise comparison, scoring, matrices
• Do 17/10 B10 Perl Herling Demo
• Fr 18/10 Multiple alignment, Basic and advanced
  Database Searching
• Do geen practicum
• Fr 25/10 geen les
• Do geen practicum
• Fr 1/11 geen les
• Do 7/11 Demo Oefeningen
• Fr 8/11 Phylogenetics, Gene prediction, junk
  mining (RNA prediction)
• Fr 15/11 geen les
• Fr 22/11 Protein structure, classification and
  engineering

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Blast

• Examples
• Proteomics, 2D analysis
• Yeast Two-Hybrid positives
• Sequencing
• Orthology Search
• .

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Blast

• BLAST is actually a family of programs
• BLASTN - Nucleotide query searching a nucleotide
  database.
• BLASTP - Protein query searching a protein
  database.
• BLASTX - Translated nucleotide query sequence (6
  frames) searching a protein database.
• TBLASTN - Protein query searching a translated
  nucleotide (6 frames) database.
• TBLASTX - Translated nucleotide query (6 frames)
  searching a translated nucleotide (6 frames)
  database.

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Oefeningen http//biochema.rug.ac.be/

• Which genes are involved in the PRADER-WILLI
  SYNDROME ?
• How may different human PDE (phosphodiesterases)
  are available in Genbank ?
• How big is the anthrax genome and how many genes
  are present ?
• Which of the 4 sequences (seq1/2/3/4)
• Contains a hexokinases signature
  (LIVM-G-F-TN-F-S-FY-P-x(5)-LIVM-DNST-x(3
  )-LIVM- x(2)-W-T-K-x-LF)
• How many of them?
• Where (hint) ?
• Write program (random.pl) to generate 10 random
  sequences of 1000 bp and write them to a file in
  fasta format
• Find the answer in ultimate-sequence.txt
• (hint use AA1 to perform translation(s))
• What is the restriction enzyme which the longest
  recognition site ?

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Pattern.pl

• We would like to find out whether the concensus
  sequence is contained (somewhere) in a given
  sequence a.
• Without quantifiers
• if (a /ACCCCAGAGAGGTGT/) ...
• With quantifiers
• if (a /AC4AG3(GT)2/) ...

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• gtSEQ1
• MGNLFENCTHRYSFEYIYENCTNTTNQCGLIRNVASSIDVFHWLDVYIST
  TIFVISGILNFYCLFIALYT YYFLDNETRKHYVFVLSRFLSSILVIISL
  LVLESTLFSESLSPTFAYYAVAFSIYDFSMDTLFFSYIMIS
  LITYFGVVHYNFYRRHVSLRSLYIILISMWTFSLAIAIPLGLYEAASNSQ
  GPIKCDLSYCGKVVEWITCS LQGCDSFYNANELLVQSIISSVETLVGSL
  VFLTDPLINIFFDKNISKMVKLQLTLGKWFIALYRFLFQMT
  NIFENCSTHYSFEKNLQKCVNASNPCQLLQKMNTAHSLMIWMGFYIPSAM
  CFLAVLVDTYCLLVTISILK SLKKQSRKQYIFVVVRLSAAILIALCIII
  IQSTYFIDIPFRDTFAFFAVLFIIYDFSILSLLGSFTGVAM
  MTYFGVMRPLVYRDKFTLKTIYIIAFAIVLFSVCVAIPFGLFQAADEIDG
  PIKCDSESCELIVKWLLFCI ACLILMGCTGTLLFVTVSLHWHSYKSKKM
  GNVSSSAFNHGKSRLTWTTTILVILCCVELIPTGLLAAFGK
  SESISDDCYDFYNANSLIFPAIVSSLETFLGSITFLLDPIINFSFDKRIS
  KVFSSQVSMFSIFFCGKR
• gtSEQ2
• MLDDRARMEA AKKEKVEQIL AEFQLQEEDL KKVMRRMQKE
  MDRGLRLETH EEASVKMLPT YVRSTPEGSE VGDFLSLDLG
  GTNFRVMLVK VGEGEEGQWS VKTKHQMYSI PEDAMTGTAE
  MLFDYISECI SDFLDKHQMK HKKLPLGFTF SFPVRHEDID
  KGILLNWTKG FKASGAEGNN VVGLLRDAIK RRGDFEMDVV
  AMVNDTVATM ISCYYEDHQC EVGMIVGTGC NACYMEEMQN
  VELVEGDEGR MCVNTEWGAF GDSGELDEFL LEYDRLVDES
  SANPGQQLYE KLIGGKYMGE LVRLVLLRLV DENLLFHGEA
  SEQLRTRGAF ETRFVSQVES DTGDRKQIYN ILSTLGLRPS
  TTDCDIVRRA CESVSTRAAH MCSAGLAGVI NRMRESRSED
  VMRITVGVDG SVYKLHPSFK ERFHASVRRL TPSCEITFIE
  SEEGSGRGAA LVSAVACKKA CMLGQ
• gtSEQ3
• MESDSFEDFLKGEDFSNYSYSSDLPPFLLDAAPCEPESLEINKYFVVIIY
  VLVFLLSLLGNSLVMLVILY SRVGRSVTDVYLLNLALADLLFALTLPIW
  AASKVTGWIFGTFLCKVVSLLKEVNFYSGILLLACISVDRY
  LAIVHATRTLTQKRYLVKFICLSIWGLSLLLALPVLIFRKTIYPPYVSPV
  CYEDMGNNTANWRMLLRILP QSFGFIVPLLIMLFCYGFTLRTLFKAHMG
  QKHRAMRVIFAVVLIFLLCWLPYNLVLLADTLMRTWVIQET
  CERRNDIDRALEATEILGILHSCLNPLIYAFIGQKFRHGLLKILAIHGLI
  SKDSLPKDSRPSFVGSSSGH TSTTL
• gtSEQ4
• MEANFQQAVK KLVNDFEYPT ESLREAVKEF DELRQKGLQK
  NGEVLAMAPA FISTLPTGAE TGDFLALDFG GTNLRVCWIQ
  LLGDGKYEMK HSKSVLPREC VRNESVKPII DFMSDHVELF
  IKEHFPSKFG CPEEEYLPMG FTFSYPANQV SITESYLLRW
  TKGLNIPEAI NKDFAQFLTE GFKARNLPIR IEAVINDTVG
  TLVTRAYTSK ESDTFMGIIF GTGTNGAYVE QMNQIPKLAG
  KCTGDHMLIN MEWGATDFSC LHSTRYDLLL DHDTPNAGRQ
  IFEKRVGGMY LGELFRRALF HLIKVYNFNE GIFPPSITDA
  WSLETSVLSR MMVERSAENV RNVLSTFKFR FRSDEEALYL
  WDAAHAIGRR AARMSAVPIA SLYLSTGRAG KKSDVGVDGS
  LVEHYPHFVD MLREALRELI GDNEKLISIG IAKDGSGIGA
  ALCALQAVKE KKGLA MEANFQQAVK KLVNDFEYPT ESLREAVKEF
  DELRQKGLQK NGEVLAMAPA FISTLPTGAE TGDFLALDFG
  GTNLRVCWIQ LLGDGKYEMK HSKSVLPREC VRNESVKPII
  DFMSDHVELF IKEHFPSKFG CPEEEYLPMG FTFSYPANQV
  SITESYLLRW TKGLNIPEAI NKDFAQFLTE GFKARNLPIR
  IEAVINDTVG TLVTRAYTSK ESDTFMGIIF GTGTNGAYVE
  QMNQIPKLAG KCTGDHMLIN MEWGATDFSC LHSTRYDLLL
  DHDTPNAGRQ IFEKRVGGMY LGELFRRALF HLIKVYNFNE
  GIFPPSITDA WSLETSVLSR MMVERSAENV RNVLSTFKFR
  FRSDEEALYL WDAAHAIGRR AARMSAVPIA SLYLSTGRAG
  KKSDVGVDGS LVEHYPHFVD MLREALRELI GDNEKLISIG
  IAKDGSGIGA ALCALQAVKE KKGLA

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Use w and strict

• !C\Perl\Bin\perl.exe
• value 42
• print "Value is OK\n" if valu lt 100
• Using my
• You can use "my" on a single variable, or on a
  list of variables my value 42 my a my
  (c,d,e,f) my (first,second) (1,2)

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Sub routine

• a5b9sumAdd(5,9)print "The SUM is
  sum\n" sub Add() a_0
• b_1
• alternatively we could do this
  my(a,b)_at__ my(answer)ab return
  answer

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Sub routine

• sub random_nucleotide
• my(_at_nucs) _at__
• return nucsrand _at_nucs
• Rand Function
• number int(rand(5)) will produce possible
  numbers of 0,1,2,3,4

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Translate.pl

• gtultimate-sequence
• ACTCGTTATGATATTTTTTTTGAACGTGAAAATACTTTTCGTGCTATGGA
  AGGACTCGTTATCGTGAAGTTGAACGTTCTGAATGTATGCCTCTTGAAAT
  GGAAAATACTCATTGTTTATCTGAAATTTGAATGGGAATTTTATCTACAA
  TGTTTTATTCTTACAGAACATTAAATTGTGTTATGTTTCATTTCACATTT
  TAGTAGTTTTTTCAGTGAAAGCTTGAAAACCACCAAGAAGAAAAGCTGGT
  ATGCGTAGCTATGTATATATAAAATTAGATTTTCCACAAAAAATGATCTG
  ATAAACCTTCTCTGTTGGCTCCAAGTATAAGTACGAAAAGAAATACGTTC
  CCAAGAATTAGCTTCATGAGTAAGAAGAAAAGCTGGTATGCGTAGCTATG
  TATATATAAAATTAGATTTTCCACAAAAAATGATCTGATAA

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• my AA1 (
• 'UUU','F',
• 'UUC','F',
• 'UUA','L',
• 'UUG','L',
• 'UCU','S',
• 'UCC','S',
• 'UCA','S',
• 'UCG','S',
• 'UAU','Y',
• 'UAC','Y',
• 'UAA','',
• 'UAG','',
• 'UGU','C',
• 'UGC','C',
• 'UGA','',
• 'UGG','W',
• 'CUU','L',

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Web application

• Install/Run Apache (PHPTriad)
• Input Form in C\apache\HTDOCS
• Eg translate.html
• Action to .pl script in C\apache\cgi-bin that
  generate HTML (using the CGI module)

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Sliding window GC, Chou-Fasman, Dotplot

• sub gc_content
• my seq shift
• my win shift
• print "length ",length(seq),"\n"
• for (my i 0 i lt length(seq) - win i)
• my segment substr(seq,i,win)
• my gc_count segment tr/GCgc/GCgc/
• print i1,"\t",segment,"\t",gc_count,"\n"

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• Combinatie Blast ClustalW
• Structure/Function - Directed Evolution
• Rather than rely upon natural variation, directed
  evolution generates new variation in one of two
  ways.
• The first is by inducing mutations in the gene of
  interest, either by using mutator strains, or by
  error-prone PCR (in which the fidelity of Taq
  polymerase is decreased rather than optimised).
• The second involves shuffling different regions
  of members of the same gene family to produce
  genes in which the inherent variation of the
  family is recombined to give novel gene products.
  
• Once a population has been produced, screening
  and selection can isolate those members which
  most closely match the desired attributes. The
  whole process can then be repeated using these
  members as the starting point for the generation
  of new variation. After relatively few cycles,
  great improvements in performance have been seen.