1 / 40

Biologia Molecular , revisão do conteúdo

Biologia Molecular , revisão do conteúdo. Prof Francisco Prosdocimi. Á CIDOS N UCLEICOS. DNA, RNA Armazenamento da informação genética Polímeros de nucleotídeos. DNA E RNA. Polímeros de nucleotídeos Esqueleto de ribose-fosfato ligado às bases nitrogenadas. R EPLICAÇÃO DO DNA.

joelle-wall
Download Presentation

Biologia Molecular , revisão do conteúdo

An Image/Link below is provided (as is) to download presentation Download Policy: Content on the Website is provided to you AS IS for your information and personal use and may not be sold / licensed / shared on other websites without getting consent from its author. Content is provided to you AS IS for your information and personal use only. Download presentation by click this link. While downloading, if for some reason you are not able to download a presentation, the publisher may have deleted the file from their server. During download, if you can't get a presentation, the file might be deleted by the publisher.

E N D

Presentation Transcript

  1. Biologia Molecular, revisão do conteúdo Prof Francisco Prosdocimi

  2. ÁCIDOS NUCLEICOS • DNA, RNA • Armazenamento da informação genética • Polímeros de nucleotídeos

  3. DNA E RNA • Polímeros de nucleotídeos • Esqueleto de ribose-fosfato ligado às bases nitrogenadas

  4. REPLICAÇÃO DO DNA • O DNA é composto por uma dupla-hélice • Replicação semi-conservativa: as bases presentes em uma das fitas contém toda a informação necessária para a síntese da nova fita • A complementaridade das bases A = T, G = C • As duas fitas do DNA são antiparalelas

  5. EVOLUÇÃO POR MUTAÇÕES • A modificação das moléculas de DNA ao longo do tempo (mutação) é um dos principais fatores evolutivos

  6. DOGMA CENTRAL E TRADUÇÃO

  7. PROTEÍNAS • Moléculas mais importantes? • Polímeros de aminoácidos • Apenas 20 diferentes aminoácidos estão presentes nas moléculas biológicas Carboxil Amino

  8. AMINOÁCIDOS

  9. LIGANDO AMINOÁCIDOS • Onde acontece? • Quem atua como catalisador?

  10. ESTRUTURA DAS PROTEÍNAS • Enovelamentode proteínas

  11. HIERARQUIA ESTRUTURAL

  12. ALFABETO QUÍMICO • Todos os organismos vivos são constituídos a partir das mesmas unidades monoméricas • A estrutura das macromoléculas é o que determina a sua função biológica • Cada espécie apresenta um conjunto distinto de macromoléculas

  13. O Sequenciamento de moléculas de DNA Prof. Dr. Francisco Prosdocimi >gi|33869444|gb|BC008730.2| Homo sapiens hexokinase 1, mRNA (cDNA clone MGC:1724 IMAGE:3163058), complete cds GGCTGCGGAGGACCGACCGTCCCCACGCCTGCCGCCCCGCGACCCCGACCGCCAGCATGATCGCCGCGCA GCTCCTGGCCTATTACTTCACGGAGCTGAAGGATGACCAGGTCAAAAAGATTGACAAGTATCTGTATGCC ATGCGGCTCTCCGATGAAACTCTCATAGATATCATGACTCGCTTCAGGAAGGAGATGAAGAATGGCCTCT CCCGGGATTTTAATCCAACAGCCACAGTCAAGATGTTGCCAACATTCGTAAGGTCCATTCCTGATGGCTC TGAAAAGGGAGATTTCATTGCCCTGGATCTTGGTGGGTCTTCCTTTCGAATTCTGCGGGTGCAAGTGAAT CATGAGAAAAACCAGAATGTTCACATGGAGTCCGAGGTTTATGACACCCCAGAGAACATCGTGCACGGCA GTGGAAGCCAGCTTTTTGATCATGTTGCTGAGTGCCTGGGAGATTTCATGGAGAAAAGGAAGATCAAGGA CAAGAAGTTACCTGTGGGATTCACGTTTTCTTTTCCTTGCCAACAATCCAAAATAGATGAGGCCATCCTG ATCACCTGGACAAAGCGATTTAAAGCGAGCGGAGTGGAAGGAGCAGATGTGGTCAAACTGCTTAACAAAG(...)TGACAGGCCTTCTGGGCCTCCAAAGCCCATCCTTGGGGTTCCCCCTCCCTGTGTGAAATGTATTATCACCAGCAGACACTGCCGGGCCTCCCTCCCGGGGGCACTGCCTGAAGGCGAGTGTGGGCATAGCATTAGCTGCT TCCTCCCCTCCTGGCACCCACTGTGGCCTGGCATCGCATCGTGGTGTGTCAATGCCACAAAATCGTGTGT CCGTGGAACCAGTCCTAGCCGCGTGTGACAGTCTTGCATTCTGTTTGTCTCGTGGGGGGAGGTGGACAGT CCTGCGGAAATGTGTCTTGTCTCCATTTGGATAAAAGGAACCAACCAACAAACAATGCCATCACTGGAAT TTCCCACCGCTTTGTGAGCCGTGTCGTATGACCTAGTAAACTTTGTACCAATTCAAAAAAAAAAAAAAAAAA

  14. Bioquímica + Biomol • Enzimas são proteínas, portanto: • São formadas por sequências de aminoácidos • Derivam de informações dispostas por genes no DNA, que deve ser transcrito e, posteriormente, traduzido • Podemos saber a sequência delas, tanto de aminoácidos quanto de nucleotídeos

  15. >gi|188497753|ref|NM_000188.2| Homo sapiens hexokinase 1 (HK1), nuclear gene encoding mitochondrial protein, transcript variant 1, mRNA GAGGAGGAGCCGCCGAGCAGCCGCCGGAGGACCACGGCTCGCCAGGGCTGCGGAGGACCGACCGTCCCCA CGCCTGCCGCCCCGCGACCCCGACCGCCAGCATGATCGCCGCGCAGCTCCTGGCCTATTACTTCACGGAG CTGAAGGATGACCAGGTCAAAAAGATTGACAAGTATCTCTATGCCATGCGGCTCTCCGATGAAACTCTCA TAGATATCATGACTCGCTTCAGGAAGGAGATGAAGAATGGCCTCTCCCGGGATTTTAATCCAACAGCCAC AGTCAAGATGTTGCCAACATTCGTAAGGTCCATTCCTGATGGCTCTGAAAAGGGAGATTTCATTGCCCTG GATCTTGGTGGGTCTTCCTTTCGAATTCTGCGGGTGCAAGTGAATCATGAGAAAAACCAGAATGTTCACA TGGAGTCCGAGGTTTATGACACCCCAGAGAACATCGTGCACGGCAGTGGAAGCCAGCTTTTTGATCATGT TGCTGAGTGCCTGGGAGATTTCATGGAGAAAAGGAAGATCAAGGACAAGAAGTTACCTGTGGGATTCACG TTTTCTTTTCCTTGCCAACAATCCAAAATAGATGAGGCCATCCTGATCACCTGGACAAAGCGATTTAAAG CGAGCGGAGTGGAAGGAGCAGATGTGGTCAAACTGCTTAACAAAGCCATCAAAAAGCGAGGGGACTATGA TGCCAACATCGTAGCTGTGGTGAATGACACAGTGGGCACCATGATGACCTGTGGCTATGACGACCAGCAC TGTGAAGTCGGCCTGATCATCGGCACTGGCACCAATGCTTGCTACATGGAGGAACTGAGGCACATTGATC TGGTGGAAGGAGACGAGGGGAGGATGTGTATCAATACAGAATGGGGAGCCTTTGGAGACGATGGATCATT AGAAGACATCCGGACAGAGTTTGACAGGGAGATAGACCGGGGATCCCTCAACCCTGGAAAACAGCTGTTT GAGAAGATGGTCAGTGGCATGTACTTGGGAGAGCTGGTTCGACTGATCCTAGTCAAGATGGCCAAGGAGG GCCTCTTATTTGAAGGGCGGATCACCCCGGAGCTGCTCACCCGAGGGAAGTTTAACACCAGTGATGTGTC AGCCATCGAAAAGAATAAGGAAGGCCTCCACAATGCCAAAGAAATCCTGACCCGCCTGGGAGTGGAGCCG TCCGATGATGACTGTGTCTCAGTCCAGCACGTTTGCACCATTGTCTCATTTCGCTCAGCCAACTTGGTGG CTGCCACACTGGGCGCCATCTTGAACCGCCTGCGTGATAACAAGGGCACACCCAGGCTGCGGACCACGGT TGGTGTCGACGGATCTCTTTACAAGACGCACCCACAGTATTCCCGGCGTTTCCACAAGACTCTAAGGCGC TTGGTGCCAGACTCCGATGTGCGCTTCCTCCTCTCGGAGAGTGGCAGCGGCAAGGGGGCTGCCATGGTGA CGGCGGTGGCCTACCGCTTGGCCGAGCAGCACCGGCAGATAGAGGAGACCCTGGCTCATTTCCACCTCAC CAAGGACATGCTGCTGGAGGTGAAGAAGAGGATGCGGGCCGAGATGGAGCTGGGGCTGAGGAAGCAGACG CACAACAATGCCGTGGTTAAGATGCTGCCCTCCTTCGTCCGGAGAACTCCCGACGGGACCGAGAATGGTG ACTTCTTGGCCCTGGATCTTGGAGGAACCAATTTCCGTGTGCTGCTGGTGAAAATCCGTAGTGGGAAAAA GAGAACGGTGGAAATGCACAACAAGATCTACGCCATTCCTATTGAAATCATGCAGGGCACTGGGGAAGAG CTGTTTGATCACATTGTCTCCTGCATCTCTGACTTCTTGGACTACATGGGGATCAAAGGCCCCAGGATGC CTCTGGGCTTCACGTTCTCATTTCCCTGCCAGCAGACGAGTCTGGACGCGGGAATCTTGATCACGTGGAC AAAGGGTTTTAAGGCAACAGACTGCGTGGGCCACGATGTAGTCACCTTACTAAGGGATGCGATAAAAAGG AGAGAGGAATTTGACCTGGACGTGGTGGCTGTGGTCAACGACACAGTGGGCACCATGATGACCTGTGCTT ATGAGGAGCCCACCTGTGAGGTTGGACTCATTGTTGGGACCGGCAGCAATGCCTGCTACATGGAGGAGAT GAAGAACGTGGAGATGGTGGAGGGGGACCAGGGGCAGATGTGCATCAACATGGAGTGGGGGGCCTTTGGG GACAACGGGTGTCTGGATGATATCAGGACACACTACGACAGACTGGTGGACGAATATTCCCTAAATGCTG GGAAACAAAGGTATGAGAAGATGATCAGTGGTATGTACCTGGGTGAAATCGTCCGCAACATCTTAATCGA CTTCACCAAGAAGGGATTCCTCTTCCGAGGGCAGATCTCTGAGACGCTGAAGACCCGGGGCATCTTTGAG ACCAAGTTTCTCTCTCAGATCGAGAGTGACCGATTAGCACTGCTCCAGGTCCGGGCTATCCTCCAGCAGC TAGGTCTGAATAGCACCTGCGATGACAGTATCCTCGTCAAGACAGTGTGCGGGGTGGTGTCCAGGAGGGC CGCACAGCTGTGTGGCGCAGGCATGGCTGCGGTTGTGGATAAGATCCGCGAGAACAGAGGACTGGACCGT CTGAATGTGACTGTGGGAGTGGACGGGACACTCTACAAGCTTCATCCACACTTCTCCAGAATCATGCACC AGACGGTGAAGGAACTGTCACCAAAATGTAACGTGTCCTTCCTCCTGTCTGAGGATGGCAGCGGCAAGGG GGCCGCCCTCATCACGGCCGTGGGCGTGCGGTTACGCACAGAGGCAAGCAGCTAAGAGTCCGGGATCCCC AGCCTACTGCCTCTCCAGCACTTCTCTCTTCAAGCGGCGACCCCCTACCCTCCCAGCGAGTTGCGCTGGG AGACGCTGGCGCCAGGGCCTGCCGGCGCGGGGAGGAAAGCAAAATCCAACTAATGGTATATATTGTAGGG TACAGAATAGAGCGTGTGCTGTTGATAATATCTCTCACCCGGATCCCTCCTCACTTGCCCTGCCACTTTG CATGGTTTGATTTTGACCTGGTCCCCCACGTGTGAAGTGTAGTGGCATCCATTTCTAATGTATGCATTCA TCCAACAGAGTTATTTATTGGCTGGAGATGGAAAATCACACCACCTGACAGGCCTTCTGGGCCTCCAAAG CCCATCCTTGGGGTTCCCCCTCCCTGTGTGAAATGTATTATCACCAGCAGACACTGCCGGGCCTCCCTCC CGGGGGCACTGCCTGAAGGCGAGTGTGGGCATAGCATTAGCTGCTTCCTCCCCTCCTGGCACCCACTGTG GCCTGGCATCGCATCGTGGTGTGTCAATGCCACAAAATCGTGTGTCCGTGGAACCAGTCCTAGCCGCGTG TGACAGTCTTGCATTCTGTTTGTCTCGTGGGGGGAGGTGGACAGTCCTGCGGAAATGTGTCTTGTCTCCA TTTGGATAAAAGGAACCAACCAACAAACAATGCCATCACTGGAATTTCCCACCGCTTTGTGAGCCGTGTC GTATGACCTAGTAAACTTTGTACCAATTCAAAAAAAAAAAAAAAAAA >gi|188497754|ref|NP_000179.2| hexokinase 1 isoform HKI [Homo sapiens] MIAAQLLAYYFTELKDDQVKKIDKYLYAMRLSDETLIDIMTRFRKEMKNGLSRDFNPTATVKMLPTFVRS IPDGSEKGDFIALDLGGSSFRILRVQVNHEKNQNVHMESEVYDTPENIVHGSGSQLFDHVAECLGDFMEK RKIKDKKLPVGFTFSFPCQQSKIDEAILITWTKRFKASGVEGADVVKLLNKAIKKRGDYDANIVAVVNDT VGTMMTCGYDDQHCEVGLIIGTGTNACYMEELRHIDLVEGDEGRMCINTEWGAFGDDGSLEDIRTEFDRE IDRGSLNPGKQLFEKMVSGMYLGELVRLILVKMAKEGLLFEGRITPELLTRGKFNTSDVSAIEKNKEGLH NAKEILTRLGVEPSDDDCVSVQHVCTIVSFRSANLVAATLGAILNRLRDNKGTPRLRTTVGVDGSLYKTH PQYSRRFHKTLRRLVPDSDVRFLLSESGSGKGAAMVTAVAYRLAEQHRQIEETLAHFHLTKDMLLEVKKR MRAEMELGLRKQTHNNAVVKMLPSFVRRTPDGTENGDFLALDLGGTNFRVLLVKIRSGKKRTVEMHNKIY AIPIEIMQGTGEELFDHIVSCISDFLDYMGIKGPRMPLGFTFSFPCQQTSLDAGILITWTKGFKATDCVG HDVVTLLRDAIKRREEFDLDVVAVVNDTVGTMMTCAYEEPTCEVGLIVGTGSNACYMEEMKNVEMVEGDQ GQMCINMEWGAFGDNGCLDDIRTHYDRLVDEYSLNAGKQRYEKMISGMYLGEIVRNILIDFTKKGFLFRG QISETLKTRGIFETKFLSQIESDRLALLQVRAILQQLGLNSTCDDSILVKTVCGVVSRRAAQLCGAGMAA VVDKIRENRGLDRLNVTVGVDGTLYKLHPHFSRIMHQTVKELSPKCNVSFLLSEDGSGKGAALITAVGVR LRTEASS 917 aminoácidos 917 x 3 = 2751 3617-2751 = 866 3617 bp3,6 kb

  16. O método de Sanger, 1975 Polimerização do DNA a ser sequenciado (molde) na presença de: DNA polimerase primer tampão dNTPs (desóxinucleotídeo) ddNTPs (didesóxinucleotídeo) O que faria um nucleotídeo que, ao invés da extremidade 3’OH, tem uma extremidade 3’H? Como acontece a síntese de moléculas de DNA? http://www.youtube.com/watch?v=Mz-4LSfecM4&feature=related (dideóxi)

  17. G C T T A T T G C C A A T C G T A C T A T C C A G G G T T A G C C A C C A G T 5’ 3’ G A ATGCTTC ||||||| TACGAAGACCGTCTAGACTTGTCACAATGACTATAACGAA 3’ 5’

  18. 5’ 3’ ATGCTTC C T T T T C G C A C A T A G T A T A T C C G A G G G T T C A G C C A C G T C A G A ||||||||| TACGAAGACCGTCTAGACTTGTCACAATGACTATAACGAA 3’ 5’

  19. 5’ 3’ ATGCTTCTG C T T T G C C T A A A T C G T C A C A T C T G G A G T T G A G C C A C C A G T G A |||||||||| TACGAAGACCGTCTAGACTTGTCACAATGACTATAACGAA 3’

  20. 5’ 3’ ATGCTTCTGGCAGATCT C T A G C T T T C T A A T C T T C G A C G G A T C G G A C T G A A G T A C C A G C ||||||||||||||||| TACGAAGACCGTCTAGACTTGTCACAATGACTATAACGAA 3’ 5’

  21. 5’ 3’ ATGCTTCTGGCAGAT A A T G A G A T T G T A T C T G G C T A T T A T C G G C C A G T C C G C A C A C C ||||||||||||||| TACGAAGACCGTCTAGACTTGTCACAATGACTATAACGAA 3’ 5’

  22. 5’ 3’ ATGCTTCTGGCAGAT A C C A T T G A T T G T A G C T A C A T T A G T C G G G T A G T C C G C A C C C A ||||||||||||||| TACGAAGACCGTCTAGACTTGTCACAATGACTATAACGAA 3’ 5’

  23. 5’ 3’ ATGCTTCTGGCAGAT A T A T G C C A T G A T G T T C A G C T G T G A A T C T C G G T A G C C C C A C A ||||||||||||||| TACGAAGACCGTCTAGACTTGTCACAATGACTATAACGAA 3’ 5’

  24. 5’ 3’ ATGCTTCTGGCAGATCTGAACAGTGTTACTGAT ATGCTTCTGGCAGATCTGAACAGTGT ATGCTTCTGGCAGATCTGAACAGTGTTACTGATATTGCTT ATGCTTCTGGCAGATCTGAACAGTGTTACTGATAT ATGCTTCTGGCAGATCTGAACAGTGTTACT ATGCTTCTGGCAGATCTGAACAGT ATGCTTCTGGCAGATCTGAACAGTGTTACTGATATTGCT ATGCTTCTGGCAGATCTGAACAGTGTTACTGATATT ATGCTTCT ATGCTTCTGGCAGATCT ATGCTTCTGGCAGATCTGAACAGTGTTACTGATATTGCTT ATGCTTCTGGCAGATCTGAACAGTGTT ATGCTTCTGGCAGAT |||||||||||||||||||||||||||||||||||||||| TACGAAGACCGTCTAGACTTGTCACAATGACTATAACGAA 3’ 5’ População de moléculas Incorporação aleatória do didesóxi Quantidade precisa entre didesóxi e desóxi

  25. ATGCTTCT ATGCTTCTGGCAGAT ATGCTTCTGGCAGATCT ATGCTTCTGGCAGATCTGAACAGT ATGCTTCTGGCAGATCTGAACAGTGT ATGCTTCTGGCAGATCTGAACAGTGTT ATGCTTCTGGCAGATCTGAACAGTGTTACT ATGCTTCTGGCAGATCTGAACAGTGTTACTGAT ATGCTTCTGGCAGATCTGAACAGTGTTACTGATAT ATGCTTCTGGCAGATCTGAACAGTGTTACTGATATT ATGCTTCTGGCAGATCTGAACAGTGTTACTGATATTGCT ATGCTTCTGGCAGATCTGAACAGTGTTACTGATATTGCTT ATGCTTCTGGCAGATCTGAACAGTGTTACTGATATTGCTT

  26. molde • polimerase • dNTPs • ddGTPs • ddATPs • ddTTPs • ddCTPs G A T C

  27. ATGCTTCT ATGCTTCTG ATGCTTCTGG ATGCTTCTGGC ATGCTTCTGGCA ATGCTTCTGGCAG ATGCTTCTGGCAGA ATGCTTCTGGCAGAT ATGCTTCTGGCAGATC ATGCTTCTGGCAGATCT ATGCTTCTGGCAGATCTG ATGCTTCTGGCAGATCTGA ATGCTTCTGGCAGATCTGAA ATGCTTCTGGCAGATCTGAAC ATGCTTCTGGCAGATCTGAACA ATGCTTCTGGCAGATCTGAACAG ATGCTTCTGGCAGATCTGAACAGT ATGCTTCTGGCAGATCTGAACAGTG ATGCTTCTGGCAGATCTGAACAGTGT ATGCTTCTGGCAGATCTGAACAGTGTT ATGCTTCTGGCAGATCTGAACAGTGTTA ATGCTTCTGGCAGATCTGAACAGTGTTAC ATGCTTCTGGCAGATCTGAACAGTGTTACT ATGCTTCTGGCAGATCTGAACAGTGTTACTG ATGCTTCTGGCAGATCTGAACAGTGTTACTGA ATGCTTCTGGCAGATCTGAACAGTGTTACTGAT ATGCTTCTGGCAGATCTGAACAGTGTTACTGATA ATGCTTCTGGCAGATCTGAACAGTGTTACTGATAT ATGCTTCTGGCAGATCTGAACAGTGTTACTGATATT ATGCTTCTGGCAGATCTGAACAGTGTTACTGATATTG ATGCTTCTGGCAGATCTGAACAGTGTTACTGATATTGC G A T C ATGCTTCTGGCAGATCTGAACAGTGTTACTGATATTGCT ATGCTTCTGGCAGATCTGAACAGTGTTACTGATATTGCTT ATGCTTCTGGCAGATCTGAACAGTGTTACTGATATTGCTT

  28. ATGCTTCT ATGCTTCTG ATGCTTCTGG ATGCTTCTGGC ATGCTTCTGGCA ATGCTTCTGGCAG ATGCTTCTGGCAGA ATGCTTCTGGCAGAT ATGCTTCTGGCAGATC ATGCTTCTGGCAGATCT ATGCTTCTGGCAGATCTG ATGCTTCTGGCAGATCTGA ATGCTTCTGGCAGATCTGAA ATGCTTCTGGCAGATCTGAAC ATGCTTCTGGCAGATCTGAACA ATGCTTCTGGCAGATCTGAACAG ATGCTTCTGGCAGATCTGAACAGT ATGCTTCTGGCAGATCTGAACAGTG ATGCTTCTGGCAGATCTGAACAGTGT ATGCTTCTGGCAGATCTGAACAGTGTT ATGCTTCTGGCAGATCTGAACAGTGTTA ATGCTTCTGGCAGATCTGAACAGTGTTAC ATGCTTCTGGCAGATCTGAACAGTGTTACT ATGCTTCTGGCAGATCTGAACAGTGTTACTG ATGCTTCTGGCAGATCTGAACAGTGTTACTGA ATGCTTCTGGCAGATCTGAACAGTGTTACTGAT ATGCTTCTGGCAGATCTGAACAGTGTTACTGATA ATGCTTCTGGCAGATCTGAACAGTGTTACTGATAT ATGCTTCTGGCAGATCTGAACAGTGTTACTGATATT ATGCTTCTGGCAGATCTGAACAGTGTTACTGATATTG ATGCTTCTGGCAGATCTGAACAGTGTTACTGATATTGC ATGCTTCTGGCAGATCTGAACAGTGTTACTGATATTGCT ATGCTTCTGGCAGATCTGAACAGTGTTACTGATATTGCTT ATGCTTCTGGCAGATCTGAACAGTGTTACTGATATTGCTT

  29. As máquinas necessárias para o sequenciamento • Primeira etapa: junta-se os reagentes em poços de placas e coloca-se na máquina de PCR para a reaçãode amplificação interrompida • Diferenças com relação ao PCR • Utilização de um só primer • Utilização dos ddNTPs • Uma vez prontas, as sequências de diferentes tamanhos contendo os didesóxi amplificados devem ser enviadas ao sequenciador de DNA mais próximo

  30. O que faz um sequenciador de DNA? • Segunda etapa: realiza a eletroforese capilar • O sequenciador executa a eletroforese em géis capilares ultra-finos • Um sensor é responsável por emitir um laser e verificar qual o comprimento de onda emitido pelo didesóxi

  31. A produção de bibliotecas de DNA e cDNA Prof. Dr. Francisco Prosdocimi Projetos Genoma e Transcriptoma

  32. O que é um genoma? Por que haplóide? • Conjunto haplóide de informações presentes no DNA de determinado organismo • Genomas bacterianos X Genomas eucarióticos • Cromossomos são formados por uma única molécula de DNA • Genoma humano: 22 pares de cromossomos autossomos + X + Y • O problema da variação – SNPs • Estudos genômicos e o método científico • A era da pesquisa científica sem hipótese

  33. Biblioteca de DNA e cDNA DNA Fragmentação Inserção em vetores Transformação mRNA Síntese de cDNA Inserção em vetores Transformação

  34. Biblioteca transcriptômica • Ou biblioteca de cDNA, DNA complementar • Purificação dos mRNAs • Oligos dT • Retrotranscrição • Clonagem

  35. Análises genômicas e transcriptômicas • Genoma: muito utilizado para produzir sequências completas do DNA de bactérias e vírus, que apresentam genoma compacto • Assim é possível saber se o organismo tem as vias bioquímicas completas e como ele deve se alimentar • Transcriptoma: classicamente utilizado em estudos de células cancerosas, onde a diferença na expressão dos genes deve mostrar porque a célula é tumoral • Comparação entre a expressão gênica em uma célula normal e o tumor • Comparações quaisquer entre dois estados celulares http://www.ncbi.nlm.nih.gov/sites/entrez?db=genome

  36. Expressão gênica

  37. Bioinformática, formatos de arquivo

  38. O formato FASTA • Fast Alignment: programa de alinhamento da década de 80 • Arquivo texto • FASTA e multi-FASTA >gi|188497753|ref|NM_000188.2| Homo sapiens hexokinase 1 (HK1), nuclear gene encoding mitochondrial protein, transcript variant 1, mRNA GAGGAGGAGCCGCCGAGCAGCCGCCGGAGGACCACGGCTCGCCAGGGCTGCGGAGGACCGACCGTCCCCA CGCCTGCCGCCCCGCGACCCCGACCGCCAGCATGATCGCCGCGCAGCTCCTGGCCTATTACTTCACGGAG CTGAAGGATGACCAGGTCAAAAAGATTGACAAGTATCTCTATGCCATGCGGCTCTCCGATGAAACTCTCA TAGATATCATGACTCGCTTCAGGAAGGAGATGAAGAATGGCCTCTCCCGGGATTTTAATCCAACAGCCAC AGTCAAGATGTTGCCAACATTCGTAAGGTCCATTCCTGATGGCTCTGAAAAGGGAGATTTCATTGCCCTG GATCTTGGTGGGTCTTCCTTTCGAATTCTGCGGGTGCAAGTGAATCATGAGAAAAACCAGAATGTTCACA TGGAGTCCGAGGTTTATGACACCCCAGAGAACATCGTGCACGGCAGTGGAAGCCAGCTTTTTGATCATGT TGCTGAGTGCCTGGGAGATTTCATGGAGAAAAGGAAGATCAAGGACAAGAAGTTACCTGTGGGATTCACG TTTTCTTTTCCTTGCCAACAATCCAAAATAGATGAGGCCATCCTGATCACCTGGACAAAGCGATTTAAAG CGAGCGGAGTGGAAGGAGCAGATGTGGTCAAACTGCTTAACAAAGCCATCAAAAAGCGAGGGGACTATGA TGCCAACATCGTAGCTGTGGTGAATGACACAGTGGGCACCATGATGACCTGTGGCTATGACGACCAGCAC TGTGAAGTCGGCCTGATCATCGGCACTGGCACCAATGCTTGCTACATGGAGGAACTGAGGCACATTGATC TGGTGGAAGGAGACGAGGGGAGGATGTGTATCAATACAGAATGGGGAGCCTTTGGAGACGATGGATCATT >gi|188497753|ref|NM_000188.2| Homo sapiens hexokinase 1 (HK1), nuclear gene encoding mitochondrial protein, transcript variant 1, mRNA CAAGGACATGCTGCTGGAGGTGAAGAAGAGGATGCGGGCCGAGATGGAGCTGGGGCTGAGGAAGCAGACG CACAACAATGCCGTGGTTAAGATGCTGCCCTCCTTCGTCCGGAGAACTCCCGACGGGACCGAGAATGGTG ACTTCTTGGCCCTGGATCTTGGAGGAACCAATTTCCGTGTGCTGCTGGTGAAAATCCGTAGTGGGAAAAA GAGAACGGTGGAAATGCACAACAAGATCTACGCCATTCCTATTGAAATCATGCAGGGCACTGGGGAAGAG CTGTTTGATCACATTGTCTCCTGCATCTCTGACTTCTTGGACTACATGGGGATCAAAGGCCCCAGGATGC CTCTGGGCTTCACGTTCTCATTTCCCTGCCAGCAGACGAGTCTGGACGCGGGAATCTTGATCACGTGGAC AAAGGGTTTTAAGGCAACAGACTGCGTGGGCCACGATGTAGTCACCTTACTAAGGGATGCGATAAAAAGG AGAGAGGAATTTGACCTGGACGTGGTGGCTGTGGTCAACGACACAGTGGGCACCATGATGACCTGTGCTT >gi|188497754|ref|NP_000179.2| hexokinase 1 isoform HKI [Homo sapiens] MIAAQLLAYYFTELKDDQVKKIDKYLYAMRLSDETLIDIMTRFRKEMKNGLSRDFNPTATVKMLPTFVRS IPDGSEKGDFIALDLGGSSFRILRVQVNHEKNQNVHMESEVYDTPENIVHGSGSQLFDHVAECLGDFMEK RKIKDKKLPVGFTFSFPCQQSKIDEAILITWTKRFKASGVEGADVVKLLNKAIKKRGDYDANIVAVVNDT VGTMMTCGYDDQHCEVGLIIGTGTNACYMEELRHIDLVEGDEGRMCINTEWGAFGDDGSLEDIRTEFDRE IDRGSLNPGKQLFEKMVSGMYLGELVRLILVKMAKEGLLFEGRITPELLTRGKFNTSDVSAIEKNKEGLH NAKEILTRLGVEPSDDDCVSVQHVCTIVSFRSANLVAATLGAILNRLRDNKGTPRLRTTVGVDGSLYKTH PQYSRRFHKTLRRLVPDSDVRFLLSESGSGKGAAMVTAVAYRLAEQHRQIEETLAHFHLTKDMLLEVKKR MRAEMELGLRKQTHNNAVVKMLPSFVRRTPDGTENGDFLALDLGGTNFRVLLVKIRSGKKRTVEMHNKIY AIPIEIMQGTGEELFDHIVSCISDFLDYMGIKGPRMPLGFTFSFPCQQTSLDAGILITWTKGFKATDCVG HDVVTLLRDAIKRREEFDLDVVAVVNDTVGTMMTCAYEEPTCEVGLIVGTGSNACYMEEMKNVEMVEGDQ GQMCINMEWGAFGDNGCLDDIRTHYDRLVDEYSLNAGKQRYEKMISGMYLGEIVRNILIDFTKKGFLFRG QISETLKTRGIFETKFLSQIESDRLALLQVRAILQQLGLNSTCDDSILVKTVCGVVSRRAAQLCGAGMAA VVDKIRENRGLDRLNVTVGVDGTLYKLHPHFSRIMHQTVKELSPKCNVSFLLSEDGSGKGAALITAVGVR LRTEASS

  39. O formato GenBank • http://www.ncbi.nlm.nih.gov • Comandos LINUX

More Related