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Exerccios de Bioinformtica Prof. Maria Lucila Hernandez Macedo e Prof. Leandro E. C. Diniz

Exercicio 01

A partir da sequncia abaixo, analise e responda:

1 acatccgcgg caacgcctcc ttggtgtcgt ccgcttccaa taacccagct tgcgtcctgc

61 acacttgtgg cttccgtgca cacattaaca actcatggtt ctagctccca gtcgccaagc

121 gttgccaagg cgttgagaga tcatctggga agtcttttac ccagaattgc tttgattcag

181 gccagctggt ttttcctgcg gtgattcgga aattcgcgaa ttcctctggt cctcatccag

241 gtgcgcggga agcaggtgcc caggagagag gggataatga agattccatg ctgatgatcc

301 caaagattga acctgcagac caagcgcaaa gtagaaactg aaagtacact gctggcggat

361 cctacggaag ttatggaaaa ggcaaagcgc agagccacgc cgtagtgtgt gccgcccccc

421 ttgggatgga tgaaactgca gtcgcggcgt gggtaagagg aaccagctgc agagatcacc

481 ctgcccaaca cagactcggc aactccgcgg aagaccaggg tcctgggagt gactatgggc

541 ggtgagagct tgctcctgct ccagttgcgg tcatcatgac tacgcccgcc tcccgcagac

601 catgttccat gtttctttta ggtatatctt tggacttcct cccctgatcc ttgttctgtt

661 gccagtagca tcatctgatt gtgatattga aggtaaagat ggcaaacaat atgagagtgt

721 tctaatggtc agcatcgatc aattattgga cagcatgaaa gaaattggta gcaattgcct

781 gaataatgaa tttaactttt ttaaaagaca tatctgtgat gctaataagg aaggtatgtt

841 tttattccgt gctgctcgca agttgaggca atttcttaaa atgaatagca ctggtgattt

901 tgatctccac ttattaaaag tttcagaagg cacaacaata ctgttgaact gcactggcca

961 ggttaaagga agaaaaccag ctgccctggg tgaagcccaa ccaacaaaga gtttggaaga

1021 aaataaatct ttaaaggaac agaaaaaact gaatgacttg tgtttcctaa agagactatt

1081 acaagagata aaaacttgtt ggaataaaat tttgatgggc actaaagaac actgaaaaat

1141 atggagtggc aatatagaaa cacgaacttt agctgcatcc tccaagaatc tatctgctta

1201 tgcagttttt cagagtggaa tgcttcctag aagttactga atgcaccatg gtcaaaacgg

1261 attagggcat ttgagaaatg catattgtat tactagaaga tgaatacaaa caatggaaac

1321 tgaatgctcc agtcaacaaa ctatttctta tatatgtgaa catttatcaa tcagtataat

1381 tctgtactga tttttgtaag acaatccatg taaggtatca gttgcaataa tacttctcaa

1441 acctgtttaa atatttcaag acattaaatc tatgaagtat ataatggttt caaagattca

1501 aaattgacat tgctttactg tcaaaataat tttatggctc actatgaatc tattatactg

1561 tattaagagt gaaaattgtc ttcttctgtg ctggagatgt tttagagtta acaatgatat

1621 atggataatg ccggtgagaa taagagagtc ataaacctta agtaagcaac agcataacaa

1681 ggtccaagat acctaaaaga gatttcaaga gatttaatta atcatgaatg tgtaacacag

1741 tgccttcaat aaatggtata gcaaatgttt tgacatgaaa aaaggacaat ttcaaaaaaa

1801 taaaataaaa taaaaataaa ttcacctagt ctaaggatgc taaaccttag tactgagtta

1861 cattgtcatt tatatagatt ataacttgtc taaataagtt tgcaatttgg gagatatatt

1921 tttaagataa taatatatgt ttacctttta attaatgaaa tatctgtatt taattttgac

1981 actatatctg tatataaaat attttcatac agcattacaa attgcttact ttggaataca

2041 tttctccttt gataaaataa atgagctatg tattaacaaa aaaaaaaaaa aaaaaaaaaa

2101 aaaaaaaaaa aaaaaa

O que fazer?

Voc recebeu uma sequncia na direo 53 de um cDNA (cpia de RNA mensageiro) desconhecido. As perguntas so:

1. A sequncia corresponde a qual gene? 2. De que espcie? 3. Quais os 12 nucleotdeos do incio da traduo? 4. Qual a sequncia de aminocidos codificada pelo RNA? (apresentar como

exemplo abaixo)

2

Dicas:

1. Faa um BLAST da sequncia no site: http://www.ncbi.nlm.nih.gov/BLAST/

Entre em: Nucleotide-nucleotide BLAST (blastn)

Coloque a sequncia no box superior

Aperte o boto virtual:

Na prxima pgina, aperte o boto e aguarde (no aperte vrias vezes, pois o tempo ser ainda mais lento)...

Logo abaixo voc encontrar a sequencia de DNA mais parecida com aquela que voc iniciou a procura. Voc ver o nome e a espcie e um valor de similaridade

(E value), que inversamente proporcional semelhana das sequncias

(sequncias idnticas do E value igual a ZERO).

2. Para obter a sequncia de aminocidos, v ao site: http://us.expasy.org/tools/dna.html

O programa ir analisar todos os possveis ORF (Open Reading Frames).

Explicando: o ribossomo l o RNA de 3 em 3 letras, podendo comear pela

primeira letra da sequncia, pela segunda ou pela terceira. Dependendo em qual

posio ele comea, a sequncia de aminocidos obtida ser diferente. Preste

ateno nos sinais Met (abreviao de Metionina) e Stop (abreviao de cdon

de trmino da traduo). A sequncia da protena (se a sequncia que voc est

analisando for o cDNA inteiro, de ponta a ponta) comea sempre com uma

Metionina e acaba com um Stop. Quanto mais longa for a sequncia de

aminocidos desde uma Met at um Stop, maiores chances de que esse seja o

ORF certo (o polipeptdeo, ou protena, codificado pela sequncia em questo).

Outra observao: em alguns casos no se sabe se a sequncia de cDNA a fita + ou a fita , por isso o programa vai liberar informao de traduo nas duas direes da sequncia dada.

Voc encontrar outra forma de visualizar qual a ORF certa no site (a ORF o box verde clique nele para ver a sequncia):

http://www.ncbi.nlm.nih.gov/gorf/gorf.html

Tem uma homepage com um exerccio muito interessante, onde voc faz a transcrio e depois a traduo, experimente!

http://learn.genetics.utah.edu/content/begin/dna/transcribe/

3

Exerccio 2

Aps sequenciamento do DNA de uma amostra qualquer de microrganismo isolado do

solo foi obtida a sequncia abaixo. Identifique o organismo a qual essa sequncia

pertence. Caso essa sequncia seja codificante, a que tipo de protena estaria

relacionada.

ATGGCCATACTTTGCAGTACTGCATTGGCTCTGGGCGCATGCGGAAGTATGGGGAAAGCGGGCGGCCCGACAATGCGTTGTCTATAGAACAAACAACGCAGATAGACAGCGCAGACGGGATTGATGCCTCCAAACTGCTCTTTTCCTC

TTCCCAAGCCGTTGTTATCGCCGGTGATTCTGTGGGGCAGAAGTGGGAGGGCGCGAAAGCAGCGGTGAAGCGGGGCGCGCCGCTGCTGGTGCGCACTGCCGATAACGCGTCGGCCATTGATTCGGAGATAAAGCGCCTCGGGGCTCA

AGACGTTATTAAGATTGACGAGCCTCAGGCCCCGGACCCGGAAATTTCCGAGGCAACATTGCCGATAAAATCTC

GCAGCTCACGCCCGAATCCCCGCTTTTTAACGGCGGCGCGTCCATCCTGGTCTCCGGGCACACCACGGCCGCTGATGTAGCCACCGCACGCGCGTCGGGGGCCAATGTGGAGTACCTGTCTTCGGGCGATGCGCGTGAAAGCTCTGCG

CTATCCGCTGATCCCGACGCTCATGTGGTTGCCCTGGGTCCAAGTTTTGCCAACAAAGAACGCTTTAATCGCCAG

GTAGAGATGATTAGCCATGGTGAGGTCCCCGGTGGCGGGCATCTCATTTTCCCCTCGCATCGCGTGGTAGCTCTCTACGGTCATCCTTCCGGCGGGGCGCTGGGAGTGCTTGGCGAGCAACCTGCTGAGGAAGCCGTAAACAGGGTGA

ATGATTTAGTGGGTAAGTATCAGGCCATTGCACCGGAAGAGAGCATGATCCCCGCCTTTGAGATCATTGCTACC

GTGGCGAGCTCGTCAGCAGGGCCGGATGGCAATTATTCCAATGAGGGGAACGTTGATGAGCTGCGCCCGTGGGT

TGAAGCGATTGGTGATGCTGGAGGCATAGCGATTCTTGATCTACAACCTGGCAGCGCAAGCTTCCTTGAACAGG

CACAACAATTTGAGGAATTGCTGAAACTACCGCACGTCGGACTGGCGATAGATCCCGAGTGGCGGCTTAAGCCG

GGGGAGAAACCCATGGAGAGGGTCGGCAGTGTTGGGGCGGGGGAAGTGAACCAGACTGCTGCGTGGCTGCGGGACCTGGTAAAAGATAACGAGCTCCCGCAGAAAGTCTTTGTTGTGCACCAATTTCAGCATCAGATGGTGCAGAAC

AGGGAAACCTTGGACACCACGGCACCGGAACTTTCGTGGGTTCTTCACGCAGATGGCCACGGAACCGCGGGCG

ATAAGTTTGCCACGTGGGATATGGTGCGGAAGAATCTGCAGCCCGAGTTCTACCTTGCGTGGAAGAACTTTATCGATGAGGATCAGCCGATGTTCACCCCCGAGCAGACGTTTAAGATCGAGCCTCGGCCTTGGTTTGTGTCCTATCA

ATAA

Exerccio 3:

Faa uma digesto in silico do fragmento amplificado pelos primers Eub-8f e 1492r do

genoma do organismo identificado no exerccio acima, utilizado as enzimas de

restrio: HaeIII, HhaI, MseI, MspI e RsaI.

Exerccio 4:

As sequncias abaixo correspondem a quais organismos?

Caso elas correspondessem a um gene, qual seria a funo e estrutura da protena

codificadas por cada sequncia.

Sequncia 1

CGAAAAATAAGCCATAGTCGGCACCATAAGCATAACCTAGCTCTGCGATTATCTCTAACATAATTAACTT

AAGCAGCCGTATTTATAAAGAAATTTCCAAAATAAAGCGAATATTCTAGAATCCCAAAACAAACTGGTTG

TTGCGGTAGGTCATTTGTTTGGCAGAAAGAAAACTCGAGAAATTTCTCTGGCCGTTATTCTCTATTCGTT

TTGTGACTCTCCCTCTTTGTACTATTGCTCTCTCACTCTGTCACACAGTAAACGGCGCACTGTTCTCGTT

GCTTCGAGAGAGCGCGCCTCGAATGTTCGCGAAAAGAGCGCCGGAGTATAAATAGAGGAGCTTCGTCGAC

GGAGAGTCAATTCTATTCAAACAAGCAAAGTGAACACATCGCTAAGCGAAAGCTAAGCAAACAAACAAGC

GCAGCTGAACAAGCTAAACAATCTGCAATAAAGTGCAAGTTAAAGTGAATCAATTAAAAGTAACCAACAA

CCAAGTAATTAAACTAAAAACTGCAACTACTGAAATCAACCAAGAAGTAATTATTGAAGACAAGAAGAGA

ACTCTGAATACTTTCAACAAGTCGTTACCGAGGAAGAAGAACTCACACACAATGCCTGCTATTGGAATCG

ATCTGGGCACCACCTACTCCTGCGTGGGTGTCTACCAACATGGCAAGGTGGAGATTATCGCCAACGACCA

GGGCAACCGCACCACGCCGTCCTACGTGGCTTTCACAGATTCGGAACGCCTCATCGGCGATCCGGCTAAG

AACCAGGTGGCCATGAACCCCAGAAACACAGTGTTTGACGCCAAGCGACTGATCGGCCGAAAATACGACG

ACCCCAAGATCGCAGAGGACATGAAGCACTGGCCTTTCAAGGTTGTAAGCGACGGCGGAAAGCCCAAGAT

CGGGGTGGAGTATAAGGGTGAGTCCAAGAGATTTGCCCCCGAGGAGATCAGCTCGATGGTACTGACCAAG

4

ATGAAGGAGACGGCGGAGGCATATCTGGGCGAGAGCATCACAGACGCAGTCATCACAGTTCCAGCCTACT

TCAACGACTCCCAGCGCCAGGCTACCAAAGACGCCGGTCACATCGCCGGCCTGAATGTGCTCCGCATCAT

CAATGAGCCCACGGCGGCAGCACTGGCCTACGGACTGGACAAGAACCTCAAGGGTGAGCGCAATGTGCTT

ATCTTCGACTTGGGCGGCGGCACCTTCGATGTCTCCATCCTGACCATCGACGAGGGATCACTGTTCGAGG

TGCGCTCCACCGCCGGAGACACACACTTGGGCGGCGAGGACTTTGACAACCGGCTAGTCACTCATCTGGC

GGACGAGTTCAAGCGCAAGTACAAGAAGGATCTGCGCTCCAACCCTCGCGCCCTACGACGCCTCAGAACA

GCAGCTGAACGGGCCAAGCGCACACTCTCCTCCAGCACGGAGGCCACCATCGAGATTGACGCACTGTTTG

AGGGCCAAGACTTCTACACCAAAGTGAGCCGCGCCAGGTTTGAGGAGCTGTGCGCGGACCTCTTCCGCAA

CACCCTGCAGCCTGTGGAGAAGGCCCTCAACGATGCCAAGATGGATAAGGGTCAGATCCACGACATCGTG

CTCGTCGGCGGATCCACTCGCATTCCCAAGGTGCAAAGTCTGCTGCAGGACTTCTTCCACGGCAAGAACC

TCAACCTATCCATCAACCCAGACGAGGCAGTTGCATACGGAGCTGCTGTGCAGGCCGCTATCCTCAGCGG

AGACCAGAGCGGCAAGATCCAGGACGTGCTGCTGGTGGACGTGGCCCCACTTTCATTGGGAATTGAGACC

GCTGGAGGTGTAATGACCAAGCTGATCGAGCGCAACTGCCGCATTCCGTGCAAGCAGACTAAGACGTTCT

CCACATACGCGGACAACCAGCCCGGAGTCTCCATTCAGGTGTATGAGGGCGAACGTGCGATGACGAAGGA

CAACAATGCATTGGGCACCTTCGATCTGTCCGGCATTCCACCTGCACCAAGGGGTGTGCCCCAGATAGAA

GTTACCTTCGACTTGGACGCCAATGGAATCCTGAACGTCAGCGCCAAGGAGATGAGCACGGGCAAGGCCA

AGAACATCACGATCAAGAACGACAAGGGACGGCTCTCGCAGGCCGAGATTGATCGCATGGTGAACGAGGC

TGAAAAGTACGCCGACGAGGACGAGAAGCATCGCCAGCGAATAACCTCTAGAAATGCCCTGGAGAGCTAC

GTCTTCAATGTGAAGCAGGCCGTGGAACAGGCACCTGCTGGCAAATTGGACGAGGCTGACAAGAACTCCG

TCTTGGACAAGTGCAACGACACTATCCGGTGGCTGGACAGCAACACCACTGCCGAGAAGGAGGAGTTCGA

CCACAAGCTGGAGGAGCTCACCCGCCACTGCTCCCCCATCATGACCAAGATGCATCAGCAGGGTGCGGGA

GCTGGAGCTGGTGGTCCGGGAGCAAACTGCGGCCAGCAGGCGGGAGGATTTGGAGGCTACTCTGGACCCA

CGGTCGAGGAGGTCGACTAAGGCCAAAGAGTCTAATTTTTGTTCATCAATGGGTTATAACATATGGGTTA

TATTATAAGTTTGTTTTAAGTTTTTGAGACTGATAAGAATGTTTCGATCGAATATTCCATAGAACAACAA

TAGTATTACCTAATTACCAAGTCTTAATTTAGCAAAAATGTTATTGCTTATAGAAAAAATAAATTATTTA

TTTGAAATTTAAAGTCAACTTGTCATTTAATGTTTTGTAGACTTTTGAAAGTCTTACGATACAATTAGTA

TCTAATATACATGGGTTCATTCTACATTCTATATTAGTGATGATTTCTTTAGCTAGTAATACATTTTAAT

TATATTCGGCTTTGATGATTTTCTGATTTTTTCCGAACGGATTTTCGTAGACCCTTTCGATCTCATAATG

GCTCATTTTATTGCGATGGACGGTCAGGAGAGCTCCACTTTTGAATTTCTGTTCGCAGACACCGCATTTG

TAGCACATAGCCGGGACATCCGGTTTGGGGAGATTTTCCAGTCTCTGTTGCAATTGGTTTTCGGGAATGC

GTTGCAG

Sequncia 2

GGTTCCAATCCTGCCTCTGCCACTTCTCAGTTGTATGCCCCAACCCAACCTGTCTGGCTCTGTCCTCCTT

AACAGAAGGACGGCCCTGGCCACGGGCCACAGCCAGCAACGCTTAAGCACCAGGGCCGGCGAGTGCCCTG

CCGTGGCACGGCTCCAGCGTCGCGCTCTCGAATTCATTTGCTTTCCTTAACGAGAGAAGGTTCCAGATGA

GGGCTGAACCCTCTTCGCCCCGCCCACGGCCCCTGAACGCTGGGGGAGGAGTGCATGGGGAGGGGCGGCC

CTCAAACGGGTCATTGCCATTAATAGAGACCTCAAACACCGCCTGCTAAAAATACCCGACTGGAGGAGCA

TAAAAGCGCAGCCGAGCCCAGCGCCCCGCACTTTTCTGAGCAGACGTCCAGAGCAGAGTCAGCCAGCATG

ACCGAGCGCCGCGTCCCCTTCTCGCTCCTGCGGGGCCCCAGCTGGGACCCCTTCCGCGACTGGTACCCGC

ATAGCCGCCTCTTCGACCAGGCCTTCGGGCTGCCCCGGCTGCCGGAGGAGTGGTCGCAGTGGTTAGGCGG

CAGCAGCTGGCCAGGCTACGTGCGCCCCCTGCCCCCCGCCGCCATCGAGAGCCCCGCAGTGGCCGCGCCC

GCCTACAGCCGCGCGCTCAGCCGGCAACTCAGCAGCGGGGTCTCGGAGATCCGGCACACTGCGGACCGCT

GGCGCGTGTCCCTGGATGTCAACCACTTCGCCCCGGACGAGCTGACGGTCAAGACCAAGGATGGCGTGGT

GGAGATCACCGGTGAGCCCCCCTGCTCCTGCAGGGGAGAGGAGGAGGCTAGCAGGGCGGGCAGGGCCGGG

GGCGTGCGGTTGAAACGGGGGTCCCGGGGGCCTGGGGAGTTAAACGTTGGCCCAGCACCGGGAAAAACAG

GACTCCTGATTCCCTTGCTCAGGAATTGGGAGTGCGGGTCGCTTCTAAGGGCGCTTTCTGCTCTGTAATC

CCAGCGCTTTGGGAGGCCGAGACGGGAGGATCGCTTGAGGCCAGGAGTTCAAGACTAGCCTGGGCAACAT

AGCGAGACGCGCCCCCCCGCCCCGACCCCGCGCCATTACAAAAAAAAAGCAAACAAAAATTTTTTTAAAG

ATCATCGATGAAGAGAGAAAATGCGCTTTTCTACAGAGTCCCCTTCCCACCCACAGCCCCATCCCCAGAT

AAGCGGGGAGTTCCCTGGCGCGGTGCCAGTTTCTAGCCGCTGAGTGGGCGTGTGCGCGGCTCCAAGTGCG

CCTGCGTACTGCTCACTCCCCAGCTCCGCGCCCTGCTCCGTTCCTCCCAAAACTCTGAATCGAAGAACTT

TCCGGAAGTTTCTGAGAGCCCAGACCGGCGGGCACGCCCCCATCCCCAACCCCCTCTGTTAATCCCTACC

AGCCTGCAGTCCTGGCTGCTTCCAAGCAGGAGGTGGGGCCTCTGGCCTAGCGGGGCCGAAAGGCAGTCCC

CTCCCCCGCAGTCTGATTTCCCTCTTCCCCCCAAAGGCAAGCACGAGGAGCGGCAGGACGAGCATGGCTA

CATCTCCCGGTGCTTCACGCGGAAATACACGTGAGTCCTGGCGCCAGGTCGGGGTGGGTGGGTGGCGTGG

GGGTGGGGTCAGGGAAGAGGGCACAGGGACCCACCCGGTGTGTAATGTAACGCTTGCCTTTCCTCTCTGC

ACGTCCAGGCTGCCCCCCGGTGTGGACCCCACCCAAGTTTCCTCCTCCCTGTCCCCTGAGGGCACACTGA

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CCGTGGAGGCCCCCATGCCCAAGCTAGCCACGCAGTCCAACGAGATCACCATCCCAGTCACCTTCGAGTC

GCGGGCCCAGCTTGGGGGCCCAGAAGCTGCAAAATCCGATGAGACTGCCGCCAAGTAAAGCCTTAGCCCG

GATGCCCACCCCTGCTGCCGCCACTGGCTGTGCCTCCCCCGCCACCTGTGTGTTCTTTTGATACATTTAT

CTTCTGTTTTTCTCAAATAAAGTTCAAAGCAACCACCTGTCACTGGCCCAGGCCCTGGTGTTTGTGGAAG

GAAGCCTCAGGCACCTGCCATTTGCTGGCTTTCAGGAGTCATCTTTGCTCAGGCCCGTGCTGGGCCATGT

GGGTACACTGGTGTAGGTTGCTGGACACAGGCTGACTCACATCCATAAAGACAGAGGTCTTAGGGCCGGG

CGCAGTGGCTCATACCTACAATCCCAGCACTTTGGGGGGTTGAAGCAGGAGGAGTGCTTGAAGCCAAGAG

TTCTAGACCAGCCTGGACAACA

Sequncia 3

AAACTTTCTGCGTCCGCCATCCTGTAGGAAGGATTTGTACACTTTAAACTCCCTCCCTGGTCTGAGTCCC

ACACTCTCACCACCCAGCACCTTCAGGAGCTGACCCTTAACAGCTTCACCCACAGGGACCCCGAAGTTGC

GTCGCCTCCGCAACAGTGTCAATAGCAGCACCAGCACTTCCCCACACCCTCCCCCTCAGGAATCCGTACT

CTCTAGCGAACCCCAGAAACCTCTGGAGAGTTCTGGACAAGGGCGGAACCCACAACTCCGATTACTCAAG

GGAGGCGGGGAAGCTCCACCAGACGCGAAACTGCTGGAAGATTCCTGGCCCCAAGGCCTCCTCCGGCTCG

CTGATTGGCCCAGCGGAGAGTGGGCGGGGCCGGTGAAGACTCCTTAAAGGCGCAGGGCGGCGAGCAGGGC

ACCAGACGCTGACAGCTACTCAGAATCAAATCTGGTTCCATCCAGAGACAAGCGAAGACAAGAGAAGCAG

AGCGAGCGGCGCGTTCCCGATCCTCGGCCAGGACCAGCCTTCCCCAGAGCATCCACGCCGCGGAGCGCAA

CCTTCCCAGGAGCATCCCTGCCGCGGAGCGCAACTTTCCCCGGAGCATCCACGCCGCGGAGCGCAGCCTT

CCAGAAGCAGAGCGCGGCGCCATGGCCAAGAACACGGCGATCGGCATCGACCTGGGCACCACCTACTCGT

GCGTGGGCGTGTTCCAGCACGGCAAGGTGGAGATCATCGCCAACGACCAGGGCAACCGCACGACCCCCAG

CTACGTGGCCTTCACCGACACCGAGCGCCTCATCGGGGACGCCGCCAAGAACCAGGTGGCGCTGAACCCG

CAGAACACCGTGTTCGACGCGAAGCGGCTGATCGGCCGCAAGTTCGGCGATGCGGTGGTGCAGTCCGACA

TGAAGCACTGGCCCTTCCAGGTGGTGAACGACGGCGACAAGCCCAAGGTGCAGGTGAACTACAAGGGCGA

GAGCCGGTCGTTCTTCCCGGAGGAGATCTCGTCCATGGTGCTGACGAAGATGAAGGAGATCGCTGAGGCG

TACCTGGGCCACCCGGTGACCAACGCGGTGATCACGGTGCCCGCCTACTTCAACGACTCTCAGCGGCAGG

CCACCAAGGACGCGGGCGTGATCGCCGGTCTAAACGTGCTGCGGATCATCAACGAGCCCACGGCGGCCGC

CATCGCCTACGGGCTGGACCGGACCGGCAAGGGCGAGCGCAACGTGCTCATCTTCGACCTGGGGGGCGGC

ACGTTCGACGTGTCCATCCTGACGATCGACGACGGCATCTTCGAGGTGAAGGCCACGGCGGGCGACACGC

ACCTGGGAGGGGAGGACTTCGACAACCGGCTGGTGAGCCACTTCGTGGAGGAGTTCAAGAGGAAGCACAA

GAAGGACATCAGCCAGAACAAGCGCGCGGTGCGGCGGCTGCGCACGGCGTGTGAGAGGGCCAAGAGGACG

CTGTCGTCCAGCACCCAGGCCAGCCTGGAGATCGACTCTCTGTTCGAGGGCATCGACTTCTACACATCCA

TCACGCGGGCGCGGTTCGAAGAGCTGTGCTCGGACCTGTTCCGCGGCACGCTGGAGCCCGTGGAGAAGGC

CCTGCGCGACGCCAAGATGGACAAGGCGCAGATCCACGACCTGGTGCTGGTGGGCGGCTCGACGCGCATC

CCCAAGGTGCAGAAGCTGCTGCAGGACTTCTTCAACGGGCGCGACCTGAACAAGAGCATCAACCCGGACG

AGGCGGTGGCCTACGGGGCGGCGGTGCAGGCGGCCATCCTGATGGGGGACAAGTCGGAGAACGTGCAGGA

CCTGCTGCTGCTGGACGTGGCGCCGCTGTCGCTGGGCCTGGAGACTGCGGGCGGCGTGATGACGGCGCTC

ATCAAGCGCAACTCCACCATCCCCACCAAGCAGACGCAGACCTTCACCACCTACTCGGACAACCAGCCCG

GGGTGCTGATCCAGGTGTACGAGGGCGAGAGGGCCATGACGCGCGACAACAACCTGCTGGGGCGCTTCGA

GCTGAGCGGCATCCCGCCGGCGCCCAGGGGCGTGCCGCAGATCGAGGTGACCTTCGACATCGACGCCAAC

GGCATCCTGAACGTCACGGCCACCGACAAGAGCACCGGCAAGGCCAACAAGATCACCATCACCAACGACA

AGGGCCGCCTGAGCAAGGAGGAGATCGAGCGCATGGTGCAGGAGGCCGAGCGCTACAAGGCCGAGGACGA

GGTGCAGCGCGACAGGGTGGCCGCCAAGAACGCGCTCGAGTCCTATGCCTTCAACATGAAGAGCGCCGTG

GAGGACGAGGGTCTCAAGGGCAAGCTCAGCGAGGCTGACAAGAAGAAGGTGCTGGACAAGTGCCAGGAGG

TCATCTCCTGGCTGGACTCCAACACGCTGGCCGACAAGGAGGAGTTCGTGCACAAGCGGGAGGAGCTGGA

GCGGGTGTGCAGCCCCATCATCAGTGGGCTGTACCAGGGTGCGGGTGCTCCTGGGGCTGGGGGCTTCGGG

GCCCAGGCGCCGCCGAAAGGAGCCTCTGGCTCAGGACCCACCATCGAGGAGGTGGATTAGAGGCCTCTGC

TGGCTCTCCCGGTGTGGTCTAGAAAACAGACTCTTTGCACTTGATAGCTGCTTGGGCACCGATTACTGTC

AAGGTTATTTAAAGTCTTCTTCATGGTTCAGTTTAAAGTTACAGTCTTTCTTAAGGTAATTGCGTTGACT

GTTAAATTTTGTATGCATATATATATATATATATATATATATATATATATATTCAAATATATTCAAAGTA

ATGTTGGGAGCAGCACTGTGCACTGTACCAGGGGATTATGTTTTATAGCTAATGATGTGTAAAGTCTAAA

GATTTTTTTGTAATTTTTATATCAGTGTTCCAGTAGCCTGGGAAGACATATAGTCTAGCTGCCCAGTTCC

CTGGAGATGGTCATCTCTAAGACAAAGTGTCTTAAACAAACGTCTTGGCACTGTGTACTACATAACTTTA

CTCTTTTGTACTTAAAACTTTATCTGCTTGTCCATGTTAAGGTTTTGTGGTATAACCAGTATGTTCTTTG

CATTTAATCTAAGTAGGTTAAAGATGGTGTATCCTTCCTGCATACATGTCTACACTGCCACCCTGTGTAC

ATTTTTTTCTTTGCATCACTACAAACTAATGAAAAAAACTTTTATGACTTAAATATTCAAAATAAAAGGT

TACAAGTATATTTTGTCTGTTTGTATGTTGGAAGGGCTAATGGATTCTGGGCTTCTGTGGATTTCTTAAG

TTTTTTTTAAGATTTATTATTATATGTGAACACATTGTAGCTATCTTCAGACACACCAGAAAAGGGCATC

AGATCTCATTACAGATGGGTGTGAGCCACCATGTGGTTCCTGGGATTTGAACTCAGGACCTTCGGAAGAG

6

TAGTCAGTGCTCTTAACTGCTGAGCTGTCTCTCCAGCCCCCGGATTTCTTAGTTTTGTGATAACTGGAAA

AGGGATTTTTTTTTGGTGGATTTTCAGTGCAGTTATGCAGGGAGTACAGGTATTTACTTTGAGGGTCGGG

CTCATTCATGGGAAAAAGTAGGGTGGTGCTGTTGTTTGGGGTCAGTGGAAGAGGGACTCAAGGGGCTATG

AGAGCTCAGCTC

Exerccio 5: exerccios de alinhamento A partir das sequncias abaixo, responda:

>seq 1

1 tgtgttcact agcaacctca aacagacacc atggtgcacc tgactcctga ggagaagtct

61 gccgttactg ccctgtgggg caaggtgaac gtggatgaag ttggtggtga ggccctgggc

121 aggctgctgg tggtctaccc ttggacccag aggttccttg agtcctttgg ggatctgtcc

181 actcctgatg ctgttatggg caaccctaag gtgaaggctc atggcaagaa agtgctcggt

241 gcctttagtg atggcctggc tcacctggac aacctcaagg gcacctttgc cacactgagt

301 gagctgcact gtgacaagct gcacgtggat cctgagaact tcaggctcct gggcaacgtg

361 ctggtctgtg tgctggccca tcactttggc aaagaattca ccccaccagt gcaggctgcc

421 tatcagaaag tggtggctgg tgtggctaat gccctggccc acaagtatca ctaagctcgc

481 tttcttgctg tccaatttct attaaaggtt cctttgttcc ctaagtccaa ctactaaact

541 gggggatatt atgaagggcc tt

>seq 2

1 aacgtggatg aagttggtgg tgaggccctg ggcaggctgc tggtggtcta cccttggacc

61 cagaggttct ttgagtcctt tggggatctg tccactcctg atgctgttat gggcaaccct

121 aaggtgaagg ctcatggcaa gaaagtgctc ggtgccttta gtgatggcct ggctcacctg

181 gacaacctca agggcacctt tgccacactg agtgagctgc actgtgacaa gctgcacgtg

241 gatcctgaga acttcaggct cctgggcaac gtgctggtct gtgtgctgga ccatcacttt

301 ggcaaagaat tcaccccacc agtgcaggct gcctatcaga aagtggtggc tggtgtggct

361 aatgccctgg cccacaagta tcactaagct cgctttcttg ctgtccaatt tctattaaag

421 gttcctttgt tccctaagtc caactactaa actgggggat attatgaagg gccttgagca

481 tctggatt

1. Faa o alinhamento das sequncias acima. 2. Classifique o tipo de alinhamento realizado? 3. Quais foram as posies que mostraram diferenas entre as duas sequncias? 4. Obtenha a sequncia protica dos genes. 5. H diferenas na regio que codifica a protena?

Exerccio 6:

Utilizando a sequncia abaixo:

ctactggtacttcgatctctggggccgtggcaccctggtcactgtctcctcagagtcttctctgtccaggcacc

1. Identifique o gene e a qual organismo ele pertence. 2. Obtenha a sequncia completa do gene no NCBI. 3. Alinhe as duas sequncias utilizando o programa CLUSTALW e verifique se houve

mismatches e gaps no alinhamento. 4. H alguma alterao na regio que codifica a protena?

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Exerccio 7:

Defina os seguintes itens e d exemplos de aplicao:

a) Comparao de sequncias; b) Alinhamento mltiplo; c) Primer; d) Busca booleana; e) Via metablica; f) Patente; g) Domnio.

Exerccio 8:

Execute os seguintes passos e registre (print screen) os resultados de cada etapa

marcada com *:

a) Escolha um gene com 100 ou mais aas e inclua sua sequncia em formato fasta na resposta;

b) Execute BlastP (NCBI BLAST) e registre: a tela de configurao da busca; a lista de resultados (aps o grfico) e o segundo e o terceiro alinhamento;

c) Interprete sucintamente o resultado (ignorando o primeiro alinhamento), confirmando ou no a identidade da protena e justificando a resposta;

d) Execute um alinhamento mltiplo com sete (7) protenas prximas entre si, de organismos diferentes, e registre o resultado do alinhamento. Comente o resultado;

e) Desenhe um par de primers para amplificar uma regio de 190 a 238 bp no gene desta protena, e registre configurao da ferramenta e as informaes dos primers

escolhidos;

f) Verifique se existem patentes relacionadas a esta protena, e descreva: g) Detalhes da busca executada; h) Nmero de patentes encontradas; i) Comente sucintamente uma destas patentes. j) Identifique o EC number desta protena; k) Classifique a protena segundo as ontologias do Gene Ontology.

Exerccio 9:

A sequencia abaixo est indicada em duas verses. A primeira sem edio (contendo

ntrons e xons) e a segunda editada (spliced). A partir de uma destas sequncias

indique:

a) Local onde comea e termina o gene.

b) Aps esta identificao da linha a, indique tambm a posio do gene completo no

clone BAC de 79.629bp anexo (ex. +50bp a + 350bp, ou -1800bp a -1350bp).

c) Em qual sentido esta o gene. Fita + ou fita -?

d) Marque no gene completo, quem so os ntrons e os xons.

e) Utilizando o clone BAC, e a localizao do gene feito no item b, identifique e

selecione a regio de 2 kb upstream ao gene. Qual a funo desta regio de 2kb

upstream ao gene?

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f) A partir da seleo da regio upstream, monte a sequncia completa entre regio de

2kb e gene. Monte tanto com o gene completo quanto com o gene editado.

g) Identifique se h regies promotoras em cada uma das sequencias (gene + upstream).

E se houver, indique suas posies na sequencia (use cores).

h) A partir das duas sequncias de genes, se tivssemos que desenhar 1 par de primers

para ser usado em PCR tempo real, qual seria a melhor estratgia para desenhar estes

primers? Justifique? E como fazer para saber se o cDNA usado no tempo real no esta

contaminado por gDNA?

> Gene completo CGTGGCTGATGTTGCCAGTTGAAATCGGTAAAGCAACTGCATCGCTGTATCCTGACGAGAATAATGATAC

ATCTCAAAATAATGTTATAAGTGCATTAGCCCCTCCGAAGGATGTGGATGATCTGCGGCTGATATCTGGA

TATGGAAATGTCAATATATTCACGTATAGTGAATTGAGAGCTGCTACCAAGAATTTCCGGCCAGATCAGG

TTCTTGGAGAGGGTGGCTTTGGGGTTGTATATAAAGGTGTTATTGATGAGAGTGTCAGGCCAGGTTCTGA

AACCATCCAAGTTGCTGTGAAAGAGCTAAAGTCAGATGGCTTGCAGGGAGACAAAGAGTGGCTGGTAATT

TCTCCCTTGTACTTTTTCAATATTTCTAAGCCAAATTTGTGATATTTTTTCTATCAGCATGAAATTTTCT

TCACTGCACCATGTATGAGTTGGCAATTGTAGTGAAAAACAAGTTGTCTGAAAAATGTTGTGGAGATTAC

ACCTAAGGTCTTATCTATTTCATCCTCTTGTATGTAGTTGAGTTAATGCTGTAAACACCCTCAACTTATA

TAAAATGAAGTCACCTACACAACAAACGGTTGCATTCTGCAGATCATGATTATGTTATTTAATGGCAATA

TCACTGATTGTATAAGTTGGAAACACTTTTATTGTTCTTTGGGAAATATATATTACATATTTGCTCTTCT

TTGACTTTAGGCTTAAAGGGATGTTCATCCCTAAAAGCAGATATCTAAAGATGTATGGGAATATGAAATT

TCTTCAAGTAACCACGGTATCTTCCAAGATGTAATTTATGTTACCTTATTTAGATATCTTATGGAAATCC

AGAGAGGCATTACTGTCAAGAGAAATCGTTGTCGGTCCCCCTTTGGCTCCATCTCAACTTAAAATTCAGG

AGTACATGACATGTGTAGTAAACTAATACCAAGTTTTGTCTTGGTCAGCTCCTCTATTATAAGATATATC

CATATATCAGTTTATTTGGAAGTCCAAGATGCTTAGTAACCTGTGACAAAACCCTATAATAGAAATTTGG

ACAGCCATATGATTACCAAAGATGGATTCCTTTTCCAATACCTCTGTGATCCATCTAGAATACTAATTAA

ATGAGTTACTTTTTGGGTTTTTCAGGCAGAAGTAAACTATCTGGGGCAACTTAGTCATCCCAATCTTGTC

AAACTTATTGGTTACTGCTGTGAAGGTGATCACAGGCTGCTAGTTTATGAGTATATGGCTTCTGGCAGCC

TTGACAAGCACCTATTCCGACGTAAGTACTTCTCTGACACGAATATCATTACAATACTTAACTAGTTATT

GTATAAAAATATTAATATCTGAGTTTGTGGAATTTAGTAAATGGTCATTTGTGGCCATCTATCCATTATA

ACTTCATTTTTCGTTTTTGACAACAGATAAATAATTTTTTAATCTACTATGAAGAAGGAAGACTTTATTA

CTAGAAACTAGATATTGCTGGGATATATATACCTTTTGTATATATGCATGGATTTGTTTATTATGCCTTT

GTATAACAAGAGCAATTAATTGTAACTACTAACTATCTTCATCATATCATTCCTTCCTGCCACTGGGCTC

TGTTAAGAGCAATGTTAGTAGTTTATAGTTGTTTATGGACTCTTTCCCCTGGAACCACTAATCTGGAATA

ACCTACTTATACATCTAGATATCTTTATGTATGAAAGAATTATGAAAAAGATCTCATCTATCCTCAAGTT

TTTTCATATATTCAAATAAATTTTATCTTATATTTTTTAATAATAACATAAATCTATCTTGTTATCCTCA

TATGTGGCTAACAATTAATAATAATTAATCCTTTATCCTTCCTAGTTACTAAGTGACAAGTAAACCGGTG

GGTAAGTAGTAAGCTAGGGGCAAAATTAGTAAGTATTGGGAATTGTACATGATGAGTTGTTGCCTTTTGC

TTATGAGCACAATTGCCCCTATCTAATTTCCCATATAATAAACATAAGTTTCTAAAATATACAATGATTA

AGTATAAGACAATACACTAAAAACAACTTTGCAACTTGGTAACTTTCTATATTCATTGTGTGTGTCTGAA

ATATTTCTACCTTCTTGACAGGGGTTTGTCTTACAATGCCATGGTCTACTCGAATGAAAATTGCCCTTGG

TGCTGCAAAAGGACTAGCCTTTCTTCATGCAGCTGAAAGATCAATCATCTATCGTGACTTCAAGACATCA

AATATCTTACTGGATGAAGTTTGTATCTCTTCCTGCACCATTGGACTGGATATTTAAATTCCCTAGAACT

ATCTCCATTATCATTATCATATTTAGCAAATACAACTCGGTTACAGATTATTAAGTCCCTATATATATTT

GTAGTCCATAAATTTTATGGCCATAACATTGGCAAAATTAGATAAATTTGTTATATGGTTAAGAGCACAA

TTACTTACATAAAAAAGATTTCATTCAGTGTGAACTCATCAATTTCTAAATAATGAGTCTGTTCCATTAA

AAAAAAATGCATACTTATTATTTGAAAAAGAAAATTGCAAATGTCCAGTATGTGAGCAACAAAAGTGGTT

ACTGAATCAATGAAAACAAGTAACTAAGGAACTCCATCGTATAATAATATTAAGGATACCCTTTTGAAGC

ATGCCCATACTGTGAAAGGTCATTTATATGTTTTTCATAACCTGAAAATATAGAAATCATGAAAACATAG

TTGTTTCTCATCAATTGTCTAATTCTTGGATTCACTTTGCAGGATTACAATGCAAAGCTCTCAGACTTTG

GCCTTGCAAAAGAGGGGCCTACAGGTGACCAAACTCACGTTTCCACTCGGGTCGTTGGTACATATGGATA

TGCAGCTCCCGAGTATATAATGACTGGTGAGCTTCTCAAACCACAATCCCAACTTTATGCAAAAGGGAGT

GCAGATAATTAAGCGCTACTAGCCACTTTCTATCGAGTGTCTCAAATGTGTAGCGATTACATGTCGTTTT

GTATATTTCTCTAGATGCCTTGCAGTAGGTGACTCTTTCTGGTTCTCTATTCCTTTTCTCTAAAGAAAAC

CCATGTTTCTGAGCAACTTTACCTCTATTTTAGGGCGTATAGCTTAAATTAATAACTTCTTTCATTTATT

CCAGGCCATTTAACTGCAAGGAGTGATGTTTACGGATTTGGAGTTGTATTGCTGGAGATGCTTTTAGGGA

GAAGGGCAATGGACAAGAGCAGGCCCAGCAGACACCAGAACCTCGTTGAGTGGGCTCGACCACTCCTGAT

CAATGGTCGGAAGTTGCTAAAGATCTTGGATCCAAGAATGGAAGGGCAATATTCTAACAGAGTTGCAACA

GATGTGGCTAGTTTAGCATATCGATGCCTGAGCCAGAACCCGAAAGGGAGGCCAACAATGAACCAAGTAG

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TCGAGTCGCTTGAGAGCCTTCAAGACCTGCCTGAGAACTGGGAAGGCATCCTGTTTCAGAGCAGTGAAGC

TGCTGTGACTCTCTATGAGGCTCCAAAAGAGATTGCGAGTGACCATTTAGAAAAGAACTCCAGCGAGAAT

GGAGAGAATGGATCAAATGTGCATGCCAAGGGAAGAAAGAAGCTTGGAAATGGCAGAAGCAACAGCGAGC

CACCGCCGGTGGAGTTCAGTCAGTACAGTCCTTCACCTGAGTCAGAGAGACATGAGCCAAGTAGAAGATC

AATCGATCATGACAGAATTCCAAGGCCACCTGCCTATTGACGTGGCTG

> gene editado CGTGGCTGATGTTGCCAGTTGAAATCGGTAAAGCAACTGCATCGCTGTATCCTGACGAGAATAATGATAC

ATCTCAAAATAATGTTATAAGTGCATTAGCCCCTCCGAAGGATGTGGATGATCTGCGGCTGATATCTGGA

TATGGAAATGTCAATATATTCACGTATAGTGAATTGAGAGCTGCTACCAAGAATTTCCGGCCAGATCAGG

TTCTTGGAGAGGGTGGCTTTGGGGTTGTATATAAAGGTGTTATTGATGAGAGTGTCAGGCCAGGTTCTGA

AACCATCCAAGTTGCTGTGAAAGAGCTAAAGTCAGATGGCTTGCAGGGAGACAAAGAGTGGCTGGCAGAA

GTAAACTATCTGGGGCAACTTAGTCATCCCAATCTTGTCAAACTTATTGGTTACTGCTGTGAAGGTGATC

ACAGGCTGCTAGTTTATGAGTATATGGCTTCTGGCAGCCTTGACAAGCACCTATTCCGACGGGTTTGTCT

TACAATGCCATGGTCTACTCGAATGAAAATTGCCCTTGGTGCTGCAAAAGGACTAGCCTTTCTTCATGCA

GCTGAAAGATCAATCATCTATCGTGACTTCAAGACATCAAATATCTTACTGGATGAAGATTACAATGCAA

AGCTCTCAGACTTTGGCCTTGCAAAAGAGGGGCCTACAGGTGACCAAACTCACGTTTCCACTCGGGTCGT

TGGTACATATGGATATGCAGCTCCCGAGTATATAATGACTGGCCATTTAACTGCAAGGAGTGATGTTTAC

GGATTTGGAGTTGTATTGCTGGAGATGCTTTTAGGGAGAAGGGCAATGGACAAGAGCAGGCCCAGCAGAC

ACCAGAACCTCGTTGAGTGGGCTCGACCACTCCTGATCAATGGTCGGAAGTTGCTAAAGATCTTGGATCC

AAGAATGGAAGGGCAATATTCTAACAGAGTTGCAACAGATGTGGCTAGTTTAGCATATCGATGCCTGAGC

CAGAACCCGAAAGGGAGGCCAACAATGAACCAAGTAGTCGAGTCGCTTGAGAGCCTTCAAGACCTGCCTG

AGAACTGGGAAGGCATCCTGTTTCAGAGCAGTGAAGCTGCTGTGACTCTCTATGAGGCTCCAAAAGAGAT

TGCGAGTGACCATTTAGAAAAGAACTCCAGCGAGAATGGAGAGAATGGATCAAATGTGCATGCCAAGGGA

AGAAAGAAGCTTGGAAATGGCAGAAGCAACAGCGAGCCACCGCCGGTGGAGTTCAGTCAGTACAGTCCTT

CACCTGAGTCAGAGAGACATGAGCCAAGTAGAAGATCAATCGATCATGACAGAATTCCAAGGCCACCTGC

CTATTGACGTGGCTG