MED-BIO 04-DNA-genoma

Sintesi del DNA
REPLICAZIONE
Sintesi RNA
TRASCRIZIONE
Sintesi delle proteine
TRADUZIONE
Crick’s Central Dogma
La relazione tra acidi nucleici ed informazione genetica è
sintetizzata dal dogma centrale della biologia
Un gene una proteina
(le eccezioni confermano la regola)
DNA
RNA
5’
3’
5’
3’
AGCT
TCGA
trascrizione
3’
AGCU
traduzione
Proteina
C-ter
N-ter
5’
• Il DNA è la molecola che consente il trasferimento
dell’informazione genetica da una cellula alle cellule figlie,
da una generazione alla successiva e nella popolazione
• Il RNA è la molecola che trasferisce, all’interno della
cellula, l’informazione genetica all’apparato che converte
questa in proteine, le vere molecole effettrici dei caratteri
fenotipici.
LA CELLULA ANIMALE
membrana nuleare
membrana plasmatica
Golgi
mitocondrio
perossisomi
lisosoma
nucleo
vescicole secretorie
reticolo endoplasmico
Golgi
nucleo
lisosoma
mitocondrio
reticolo endoplasmico
Geni e Genoma
L’informazione genetica presente sul DNA viene utilizzata per
dirigere la sintesi di proteine.
L’insieme delle sequenze nucleotidiche presenti nel DNA
cellulare formano il genoma umano.
La sequenza di nucleotidi sufficiente per la sintesi di una data
proteina costituisce il gene per la proteina.
Le sequenze nucleotidiche che codificano per proteine
rappresentano solo il 1-2% dell’intero genoma.
Il genoma umano è diploide – esistono due copie di DNA doppia
elica con sequenze nucleotidiche tra loro omologhe.
8
3
Il genoma umano è diploide, è organizzato in
23 coppie di cromosomi (46 cromosomi totali).
I cromosomi umani - cariotipo
E 8-2; 4th 4-10
4th 4-12
Cromosomi
Cromosomi
Struttura di un
cromosoma
eucariotico
Festival di San Remo 1999
Renato Dulbecco Premio Nobel 1975
Promotore del progetto GENOMA UMANO
Bold Words from Francis Collins:
“The history of biology was forever altered a decade ago by
the bold decision to launch a research program that would
characterize in ultimate detail the complete set of
genetic instructions of the human being.”
Francis S. Collins
Director of the National Human Genome Research Institute
N Engl J Med 1999 882:42-65
2001 la prima sequenza
“completa” del nostro genoma
Genoma umano: 6.4 x109 paia di basi
Che cosa significa avere 6.4x109 bp di informazione nel genoma ?
1 pagina di libro = 1000 lettere
1 libro = 1000 pagine x 1000 lettere/pagina
= 1.000.000 (106) lettere
6400 libri x 106 lettere/libro
6.4x109 lettere (genoma umano)
AGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGA
CAGACGACTACGACAGACTACGACAGACTACAGACTACACCCCCCCCCCCCCCCCCCCCGGGGGGGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGAC
TAAACGGGCATACGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTTTTTTTTTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAAAAAAAGCTA
CGACATACGCCAGACATACAGACTACGCTACAGCATACAGCATACGACATACGACATACCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGGGGACGACTAC
GACATACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATG
CGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACACCCCCCCAAAAAAAAAGCTACGACATACGCCAGACATACAGACTACGCTACAGCATACAGCATACGACATACG
ACATACCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGGGGACGACTACGACATACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGAC
GACTACGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACACCCCCC
GGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTCCCCCCCCCCCCCGGGGGGGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGA
ATACGACTAAACGGGCATACGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTTTTTTTTCTAGCAGATACGCATACAGACTAACGCATACGACATACGCAGCTTTTGC
AAAAGCATAGACTACGACTCAGACGCATCAGCACAAAGACGATGATTGACGTAGCATGACGATTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAAAAAAAGCTAC
GACATACGCCAGACATACAGACTACGCTACAGCATACAGCATACGACATACGACATACCCCCCCCCGATGACATACGACATACCCCCCACAAGTACGATACAGATACGACAACAACAAT
CAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGAC
TACAGACTACACCCCCCCCCCCCCCCCCCCCGGGGGGGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGCATACGAACTCAGACGACGA
CGACTACACAGCGACTAACGACGCATATTTTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAAGCTACGACATACGCCAGACATACAGACTACGCTACAGCATACA
GCATACGACATACGACATACCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGGGGACGACTACGACATACAAGTACGATACAGATACGACAACAACAATCAAT
GCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACA
GACTACACCCCCCCCCCCCGGGGGAGTAGGGTAGGCACGACGACTAGCAGACGGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGCATA
CGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTTTTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAAAAGCTACGACATACGCCAGACATACA
GACTACGCTACAGCATACAGCATACGACATACGACATACCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGGGGACGACTACGACATACAAGTACGATACAG
ATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACG
ACAGACTACGACAGACTACAGACTACACCCCCCCCCCCCCCCCCGGGGGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGCATACGAAC
TCAGACGACGACGACTACACAGCGACTAACGACGCATATTTTTTTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAAAAAAAGCTACGACATACGCCAGACATACA
GACTACGCTACAGCATACAGCATACGACATACGACATACCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGGGGACGACTACGACATACAAGTACGATACAG
ATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACG
ACAGACTACGACAGACTACAGACTACACCCCCCCCCCCCCCCCCCCCGGGGGGGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGCATA
CGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAGCTACGACATACGCCAGACATACAGACTAC
GCTACAGCATACAGCATACGACATACGACATACCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGGGGACGACTACGACATACAAGTACGATACAGATACGA
CAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGAC
TACGACAGACTACAGACTACACCCCCCCCCCCCCCCCCCCCGGGGGGGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGCATACGAACT
CAGACGACGACGACTACACAGCGACTAACGACGCATATTTTTTTTTTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAAAAAAAGCTACGACATACGCCAGACATA
CAGACTACGCTACAGCATACAGCATACGACATACGACATACCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGGGGACGACTACGACATACAAGTACGATAC
AGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTA
CGACAGACTACGACAGACTACAGACTACACCCCCCCCCCCCCCCCCCCCGGGGGGGGAAAAAGCTACGACATACGCCAGACATACAGACTACGCTACAGCATACAGCATACGACATAC
GACATACCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGGGCCCCCCCATAGCATGGGGGGGACGACTACGACATACCATGACTACCAGCATACGACATCAT
CAGCAGACGACGACGACAGCAGCAGCAGCAGACTTCGACAGCATTTGATACGACTACGACGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGG
CATACGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAATACATGACGACTGATACTGACAAT
CAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACACCCCCCGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGC
ATACGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTTTGCAAAAGCATAGACTACGACTCAGACGCATAAAAGCTACGACATACGCCAGACATACAGACTACGCTAC
AGCATACAGCATACGACATACGACATACCCCCCCCCGATGACATACGACATACCATAGCATGGGGGGGACGACTACGAAAAAAAAAAGCTACGACATACGCCAGACATACAGACTACG
CTACAGCATACAGCATACGACATACGACATACCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGCATGACGACTACGACATACAAGTGCGCGGCGCGTTGTG
TGTCGCGCGCGCTGGTAACGACAGCGCAGCAGCGCAGACACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACAAGT
ACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACACCCCCCGGGGGGGGTTACAGATACGCATACGACTACAGACTA
CGACTACGAATACGACTAAACGGGCATACGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTTTCAGCACAAAAGGGACGACTACGACATACCATGACTACCAGCATA
CGACATCATCAGCAGACGACGACGACAGCAGCAGCAGCAGACTTCGACAGCATTTGATACGACTACGACTACATGACGACTGATACTGAAGACGACTACGACAGACTACGACAGACTA
CAGACTACACCCCCCCCCCCCCCCCCCCCGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGCATACGAACTCAGACGACGACGACTACACAGCGA
CTAACGACGCATATTTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAACGGGCATACGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTTTTTTT
TTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAAAAAAAGCTACGACACAGACGACTACGACAGACTACGACAGACTACAGACTACACCCCCCCCCCCCCCCCCC
CCGGGGGGGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGCATACGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTT
TTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAAAAAAAGCTACGACATACGCCAGACATACAGACTACGCTACAGCATACAGCATACGACATACGACATACCCCCC
CCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGGGGACGACTACGACATACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAG
ACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACACCCCCCGGGGGGGGTT
ACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGCATACGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTTTTTTTTTTTGCAAAAGCATA
GACTACGACTCAGACGCATCAGCACAAAAAAAAAAAAAGCTACGACATCAGACGCATCAGCACAAAAAAACGGGCATACGAACTCAGGACTACGACAGACTACAGACTACACC
PAGINA 1
AGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGA
CAGACGACTACGACAGACTACGACAGACTACAGACTACACCCCCCCCCCCCCCCCCCCCGGGGGGGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGAC
TAAACGGGCATACGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTTTTTATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACACCCCC
CGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTCCCCCCCCCCCCCGGGGGGGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACG
AATACGACTAAACGGGCATACGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTTTTTTTTCTAGCAGATACGCATACAGACTAACGCATACGACATACGCAGCTTTTG
CAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAGACGATGATTGACGTAGCATGACGATTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAAAAAAAGCTA
CGACATACGCCAGACATACAGACTACGCTACAGCATACAGCATACGACATACGACATACCCCCCCCCGATGACATACGACATACCCCCCACAAGTACGATACAGATACGACAACAACAA
TCAATGCGATAGACAGACGACTACGACAGACTAACGACGCATATTTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAACGGGCATACGAACTCAGACGACGACGAC
TACACAGCGACTAACGACGCATATTTTTTTTTTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAAAAAAAGCTACGACACAGACGACTACGACAGACTACGACAGA
CTACAGACTACACCCCCCCCCCCCCCCCCCCCGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGA
CAGACTACAGACTACACCCCCCCCCCCCCCCCCCCCGGGGGGGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGCATACGAACTCAGAC
GACGACGACTACACAGCGACTAACGACGCATATTTTTTGCAAAAGCATAGACTACGACTCCTACGACAGACTACGACAGACTACAGACTACACCCCCCCATCAATGCGATAGACAGACG
AAAAATACATGACGACTGATACTGACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACACCCCCCGGGGGGGGTTACAGATACGCATACGACTACAGA
CTACGACTACGAATACGACTAAACGGGCATACGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTTTGCAAAAGCATAGACTACGACTCAGACGCACGACTACGACAG
ACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACACCCCCCCCCCCCGGGG
GAGTAGGGTAGGCACGACGACTAGCAGACGGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGCATACGAACTCAGACGACGACGACTAC
ACAGCGACTAACGACGCATATTTTTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAAAAGCTACGACATACGCCAGACATACAGACTACGCTACAGCATACAGCAT
ACGACATACGACATACCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGGGGACGACTACGACATACAAGTACGATACAGATACGACAACAACAATCAATGCG
ATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGAC
TACACCCCCCCCCCCCCCCCCGGGGGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGCATACGAACTCAGACGACGACGACTACACAGC
GACTAACGACGCATATTTTTTTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAAAAAAAGCTACGACATACGCCAGACATACAGACTACGCTACAGCATACAGCAT
ACGACATACGACATACCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGGGGACGACTACGACATACAAGTACGATACAGATACGACAACAACAATCAATGCG
ATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGAC
TACACCCCCCCCCCCCCCCCCCCCGGGGGGGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGCATACGAACTCAGACGACGACGACTAC
ACAGCGACTAACGACGCATATTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAGCTACGACATACGCCAGACATACAGACTACGCTACAGCATACAGCATACGACA
TACGACATACCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGGGGACGACTACGACATACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGAC
AGACGACTACGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACACC
CCCCCCCCCCCCCCCCCCGGGGGGGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGCATACGAACTCAGACGACGACGACTACACAGCG
ACTAACGACGCATATTTTTTTTTTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAAAAAAAGCTACGACATACGCCAGACATACAGACTACGCTACAGCATACAGCA
TACGACATACGACATACCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGGGGACGACTACGACATACAAGTACGATACAGATACGACAACAACAATCAATGC
GATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGA
CTACACCCCCCCCCCCCCCCCCCCCGGGGGGGGAAAAAGCTACGACATACGCCAGACATACAGACTACGCTACAGCATACAGCATACGACATACGACATACCCCCCCCCGATGACATA
CGACATACCCCCCCCCCCCCATAGCATGGGGGGCCCCCCCATAGCATGGGGGGGACGACTACGACATACCATGACTACCAGCATACGACATCATCAGCAGACGACGACGACAGCAGC
AGCAGCAGACTTCGACAGCATTTGATACGACTACGACGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGCATACGAACTCAGACGACGACGA
CTACACAGCGACTAACGACGCATATTTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAGACGCATCAGCACAAAAAAAAGCTACGACATACGCCAGACATACAGACTA
CGCTACAGCATACAGCATACGACATACGACATACCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGGGGACGACTACGACATACAAGTACGATACAGATACG
ACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGAAAAATACATGA
CGACTGATACTGACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACACCCCCCGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACG
AATACGACTAAACGGGCATACGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTTTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAAAAAAAG
CTACGACATACGCCAGACATACAGACTACGCTACAGCATACAGCATACGACATACGACATACCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGGGGACGAC
TACGACATACAAGTACGATACAGATACGACACAAAAAGACGCATCAGCACAAAAAAAAGCTACGACATACGCCAGACATACAGACTACGCTACAGCATACAGCATACGACATACGACATA
CCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGGGGACGACTACGACATACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTA
CGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAAAAAAAAAAGCTACGACATACGCCAGACATACAGACTACGCTACAGCATACAGCATACGACATA
CGACATACCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGGGGACGACTACGACATACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAG
ACGACTACGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACATTTTGCAAAAGCATAGACTACGACTCAGACGCATAAAAGCTACGACATACGCCAGACA
TACAGACTACGCTACAGCATACAGCATACGACATACGACATACCCCCCCCCGATGACATACGACATACCATAGCATGGGGGGGACGACTACGAAAAAAAAAAGCTACGACATACGCCAG
ACATACAGACTACGCTACAGCATACAGCATACGACATACGACATACCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGCATGACGACTACGACATACAAGTGC
GCGGCGCGTTGTGTGTCGCGCGCGCTGGTAACGACAGCGCAGCAGCGCAGACACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGAC
TACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACACCCCCCGGGGGGGGTTACAGATACGCATA
CGACTACAGACTACGACTACGAATACGACTAAACGGGCATACGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTTTCAGCACAAAAGGGACGACTACGACATACCAT
GACTACCAGCATACGACATCATCAGCAGACGACGACGACAGCAGCAGCAGCAGACTTCGACAGCATTTGATACGACTACGACTACATGACGACTGATACTGAAGACGACTACGACAGA
CTACGACAGACTACAGACTACCATCAGCACAAAAAAACGGGCATACGAACTCAGGACTACGACAGACTACAGACTACACC
PAGINA 2
AGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGA
CAGACGACTACGACAGACAGACTACGCTACAGCATACAGCATACGACATACGACATACCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGGGGACGACTACG
ACATACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGC
GATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACACCCCCCCAAAAAAAAAGCTACGACATACGCCAGACATACAGACTACGCTACAGCATACAGCATACGACATACGA
CATACCCCCCCCCGATGACATACATCAGCACAAAGACGATGATTGACGTAGCATGACGATTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAAAAAAAGCTACGAC
ATACGCCAGACATACAGACTACGCTACAGCATACAGCATACGACATACGACATACCCCCCCCCGATGACATACGACATACCCCCCACAAGTACGATACAGATACGACAACAACAATCAAT
GCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACATACAAGTACGATACAGATACGACAACAACA
ATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAG
ACTACAGACTACACCCCCCGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTCCCCCCCCCCCCCGGGGGGGGGGGGGGTTACAGATACGCATACGA
CTACAGACTACGACTACGAATACGACTAAACGGGCATACGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTGGGACAGACGACTACGACAGACTACGACAGACTAC
AGACTACACCCCCCCCCCCCCCCCCCCCAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAAGCTACGACATACGCCAGACATACAGACTACGCTACAGCATACAGCATACGA
CATACGACATACCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGGGGACGACTACGACATACAAGTACGATACAGATACGACAACAACAATCAATGCGATAG
ACAGACGACTACGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACA
CCCCCCCCCCCCGGGGGAGTAGGGTAGGCACGACGACTAGCAGACGGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGCATACGAACTC
AGACGACGACGACTACACAGCGACTAACGACGCATATTTTTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAAAAGCTACGACATACGCCAGACATACAGACTAC
GCTACAGCATACAGCATACGACATACGACATACCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGGGGACGACTACGACATACAAGTACGATACAGATACGA
CAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGAC
TACGACAGACTACAGACTACACCCCCCCCCCCCCCCCCGGGGGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGCATACGAACTCAGAC
GACGACGACTACACAGCGACTAACGACGCATATTTTTTTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAAAAAAAGCTACGACATACGCCAGACATACAGACTAC
GCTACAGCATACAGCATACGACATACGACATACCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGGGGACGACTACGACATACAAGTACGATACAGATACGA
CAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGAC
TACGACAGACTACAGACTACACCCCCCCCCCCCCCCCCCCCGGGGGGGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGCATACGAACT
CAGACGACGACGACTACACAGCGACTAACGACGCATATTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAGCTACGACATACGCCAGACATACAGACTACGCTACA
GCATACAGCATACGACATACGACATACCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGGGGACGACTACGACATACAAGTACGATACAGATACGACAACAA
CAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGAC
AGACTACAGACTACACCCCCCCCCCCCCCCCCCCCGGGGGGGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGCATACGAACTCAGACG
ACGACGACTACACAGCGACTAACGACGCATATTTCTACGACAGACTACAGACTACACCCCCCCCCCCCCCCCCCCCGGGGGGGGGGGGGGTTACAGATACGCATACGACTACAGACT
ACGACTACGAATACGACTAAACGGGCATACGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTTTTTTTTTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCAC
AAAAAAAAAAAAAGCTACGACATACGCCAGACATATTTTACGACGCATATTTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAATACATGACGACTGATACTGACAA
TCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACACCCCCCGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGG
CATACGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTTTGCAAAAGCATAGACTACGACTCAGACGCATAAAAGCTACGACATACGCCAGACATACAGACTACGCTA
CAGCATACAGCATACGACATACGACATACCCCCCCCCGATGACATACGAGACATACCCCCCCCCCCCCATAGCATGGGGGGGACGACTACGACATACAAGTACGATACAGATACGACA
ACAACAATCAATGCGATAGACAGACGACTACGACAGACTATTTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAAAAAAAGCTACGACATACGCCAGACATACAGA
CTACGCTACAGCATACAGCATACGACATACGACATACCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGGGGACGACTACGACATACAAGTACGATACAGAT
ACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGAC
AGACTACGACAGACTACAGACTACACCCCCCCCCCCCCCCCCCCCGGGGGGGGAAAAAGCTACGACATACGCCAGACATACAGACTACGCTACAGCATACAGCATACGACATACGACA
TACCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGGGCCCCCCCATAGCATGGGGGGGACGACTACGACATACCATGACTACCAGCATACGACATCATCAG
CAGACGACGACGACAGCAGCAGCAGCAGACTTCGACAGCATTTGATACGACTACGACGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGCAT
ACGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAATACATGACGACTGATACTGACAATCAA
TGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACACCCCCCGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGCATA
CGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTTTGCAAAAGCATAGACTACGACTCAGACGCATAAAAGCTACGACATACGCCAGACATACAGACTACGCTACAGC
ATACAGCATACGACATACGACATACCCCCCCCCGATGACATACGAGACATACCCCCCCCCCCCCATAGCATGGGGGGGACGACTACGGGGGGGGGGGGTTACAGATACGCATACGAC
TACAGACTACGACTACGAATACGACTAAACGGGCATACGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTTTTTGCAATTTTTTTCTAGCAGATACGCATACAGACTA
ACGCATACGACATACGCAGCTTTTGCAAAAGCATAGACTACGACTCAGACGCCATACCATAGCATGGGGGGGACGACTACGAAAAAAAAAAGCTACGACATACGCCAGACATACAGACT
ACGCTACAGCATACAGCATACGACATACGACATACCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGCATGACGACTACGACATACAAGTGCGCGGCGCGTT
GTGTGTCGCGCGCGCTGGTAACGACAGCGCAGCAGCGCAGACACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACA
AGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACACCCCCCGGGGGGGGTTACAGATACGCATACGACTACAGA
CTACGACTACGAATACGACTAAAAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGCATACGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATAT
TTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAAAAAAAGCTACGACATACGCCAGACATACAGACTACGCTACAGCATACAGCATACGACATACGACATACCCCC
CCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGGGGACGACTACGACATACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACA
GACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACACCCCCCGGGGGGGGT
TACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGCATACGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTTTTTTTTTTTGCAAAAGCAT
AGACTACGACTCAGACGCATCAGCACAAAAAAAAAAAAAGCTACGACATCAGACGCATCAGCACAAAAAAACGGGCATACGAACTCAGGACTACGACAGACTACAGACTACACC
PAGINA 1000
AGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGA
CAGACGACTACGACAGACTACGACAGACTACAGACTACACCCCCCCCCCCCCCCCCCCCGGGGGGGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGAC
TAAACGGGCATACGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTTTTTTTTTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAAAAAAAGCTA
CGACATACGCCAGACATACAGACTACGCTACAGCATACAGCATACGACATACGACATACCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGGGGACGACTAC
GACATACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATG
CGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACACCCCCCCAAAAAAAAAGCTACGACATACGCCAGACATACAGACTACGCTACAGCATACAGCATACGACATACG
ACATACCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGGGGACGACTACGACATACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGAC
GACTACGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACACCCCCC
GGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTCCCCCCCCCCCCCGGGGGGGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGA
ATACGACTAAACGGGCATACGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTTTTTTTTCTAGCAGATACGCATACAGACTAACGCATACGACATACGCAGCTTTTGC
AAAAGCATAGACTACGACTCAGACGCATCAGCACAAAGACGATGATTGACGTAGCATGACGATTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAAAAAAAGCTAC
GACATACGCCAGACATACAGACTACGCTACAGCATACAGCATACGACATACGACATACCCCCCCCCGATGACATACGACATACCCCCCACAAGTACGATACAGATACGACAACAACAAT
CAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGAC
TACAGACTACACCCCCCCCCCCCCCCCCCCCGGGGGGGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGCATACGAACTCAGACGACGA
CGACTACACAGCGACTAACGACGCATATTTTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAAGCTACGACATACGCCAGACATACAGACTACGCTACAGCATACA
GCATACGACATACGACATACCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGGGGACGACTACGACATACAAGTACGATACAGATACGACAACAACAATCAAT
GCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACA
GACTACACCCCCCCCCCCCGGGGGAGTAGGGTAGGCACGACGACTAGCAGACGGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGCATA
CGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTTTTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAAAAGCTACGACATACGCCAGACATACA
GACTACGCTACAGCATACAGCATACGACATACGACATACCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGGGGACGACTACGACATACAAGTACGATACAG
ATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACG
ACAGACTACGACAGACTACAGACTACACCCCCCCCCCCCCCCCCGGGGGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGCATACGAAC
TCAGACGACGACGACTACACAGCGACTAACGACGCATATTTTTTTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAAAAAAAGCTACGACATACGCCAGACATACA
GACTACGCTACAGCATACAGCATACGACATACGACATACCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGGGGACGACTACGACATACAAGTACGATACAG
ATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACG
ACAGACTACGACAGACTACAGACTACACCCCCCCCCCCCCCCCCCCCGGGGGGGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGCATA
CGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAGCTACGACATACGCCAGACATACAGACTAC
GCTACAGCATACAGCATACGACATACGACATACCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGGGGACGACTACGACATACAAGTACGATACAGATACGA
CAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGAC
TACGACAGACTACAGACTACACCCCCCCCCCCCCCCCCCCCGGGGGGGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGCATACGAACT
CAGACGACGACGACTACACAGCGACTAACGACGCATATTTTTTTTTTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAAAAAAAGCTACGACATACGCCAGACATA
CAGACTACGCTACAGCATACAGCATACGACATACGACATACCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGGGGACGACTACGACATACAAGTACGATAC
AGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTA
CGACAGACTACGACAGACTACAGACTACACCCCCCCCCCCCCCCCCCCCGGGGGGGGAAAAAGCTACGACATACGCCAGACATACAGACTACGCTACAGCATACAGCATACGACATAC
GACATACCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGGGCCCCCCCATAGCATGGGGGGGACGACTACGACATACCATGACTACCAGCATACGACATCAT
CAGCAGACGACGACGACAGCAGCAGCAGCAGACTTCGACAGCATTTGATACGACTACGACGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGG
CATACGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAATACATGACGACTGATACTGACAAT
CAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACACCCCCCGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGC
ATACGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTTTGCAAAAGCATAGACTACGACTCAGACGCATAAAAGCTACGACATACGCCAGACATACAGACTACGCTAC
AGCATACAGCATACGACATACGACATACCCCCCCCCGATGACATACGACATACCATAGCATGGGGGGGACGACTACGAAAAAAAAAAGCTACGACATACGCCAGACATACAGACTACG
CTACAGCATACAGCATACGACATACGACATACCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGCATGACGACTACGACATACAAGTGCGCGGCGCGTTGTG
TGTCGCGCGCGCTGGTAACGACAGCGCAGCAGCGCAGACACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACAAGT
ACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACACCCCCCGGGGGGGGTTACAGATACGCATACGACTACAGACTA
CGACTACGAATACGACTAAACGGGCATACGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTTTCAGCACAAAAGGGACGACTACGACATACCATGACTACCAGCATA
CGACATCATCAGCAGACGACGACGACAGCAGCAGCAGCAGACTTCGACAGCATTTGATACGACTACGACTACATGACGACTGATACTGAAGACGACTACGACAGACTACGACAGACTA
CAGACTACACCCCCCCCCCCCCCCCCCCCGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGCATACGAACTCAGACGACGACGACTACACAGCGA
CTAACGACGCATATTTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAACGGGCATACGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTTTTTTT
TTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAAAAAAAGCTACGACACAGACGACTACGACAGACTACGACAGACTACAGACTACACCCCCCCCCCCCCCCCCC
CCGGGGGGGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGCATACGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTT
TTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAAAAAAAGCTACGACATACGCCAGACATACAGACTACGCTACAGCATACAGCATACGACATACGACATACCCCCC
CCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGGGGACGACTACGACATACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAG
ACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACACCCCCCGGGGGGGGTT
ACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGCATACGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTTTTTTTTTTTGCAAAAGCATA
GACTACGACTCAGACGCATCAGCACAAAAAAAAAAAAAGCTACGACATCAGACGCATCAGCACAAAAAAACGGGCATACGAACTCAGGACTACGACAGACTACAGACTACACC
PAGINA 1 MILIONE
AGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGA
CAGACGACTACGACAGACTACGACAGACTACAGACTACACCCCCCCCCCCCCCCCCCCCGGGGGGGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGAC
TAAACGGGCATACGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTTTTTTCCCGGGGGGGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAAT
ACGACTAAACGGGCATACGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAAAAAAAGCTAC
GACATACGCCAGACATACAGACTACGCTACAGCATACAGCATACGACATACGACATACCCCCCTTTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAAAAAAAGCT
ACGACATACGCCAGACATACAGACTACGCTACAGCATACAGCATACGACATACGACATACCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGGGGACGACTA
CGACATACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAAT
GCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACACCCCCCCAAAAAAAAAGCTACGACATACGCCAGACATACAGACTACGCTACAGCATACAGCATACGACATAC
GACATACCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGGGGACGACTACGACATACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGA
CGACTACGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACACCCCC
CGGGGGGGGCCCCCCCCCCGGGGGAGTAGGGTAGGCACGACGACTAGCAGACGGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGCAT
ACGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTTTTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAAAAGCTACGACATACGCCAGACATAC
AGACTACGCTACAGCATACAGCATACGACATACGACATACCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGGGGACGACTACGACATACAAGTACGATACA
GATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTAC
GACAGACTACGACAGACTACAGACTACACCCCCCCCCCCCCCCCCGGGGGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGCATACGAA
CTCAGACGACGACGACTACACAGCGACTAACGACGCATATTTTTTTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAAAAAAAGCTACGACATACGCCAGACATAC
AGACTACGCTACAGCATACAGCATACGACATACGACATACCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGGGGACGACTACGACATACAAGTACGATACA
GATACGACAACAACAATCAATGCGATAGACAGACGACTACGACATACGCCAGACATACAGACTACGCTACAGCATACAGCATACGACATACGACATACCCCCCCCCGATGACATACGAC
ATACCCCCCCCCCCCCATAGCATGGGGGGGACGACTACGACATACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACA
GACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACACCCCCCCCCCCCCCCCCCCCGGGGGGGGGGG
GGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGCATACGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTTTTTTTTTTTGCAAAA
GCATAGACTACGACTCAGACGCATCAGCACAAAAAAAAAAAAAGCTACGACATACGCCAGACATACAGACTACGCTACAGCATACAGCATACGACATACGACATACCCCCCCCCGATGA
CATACGACATACCCCCCCCCCCCCATAGCATGGGGGGGACGACTACGACATACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGCTAAACGGGCATACGAACTCAGACG
ACCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAAGCTACGACATACGCCAGACATACAGACTACGCTACAGCATACAGCATACGACATACGACATACCCCCCCCCGAT
GACATACGACATACCCCCCCCCCCCCATAGCATGGGGGGGACGACTACGACATACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACG
ACAGACTACAGACTACCCCGGGGGGGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGCATACGAACTCAGACGACGACGACTACACAGC
GACTAACGACGCATATTTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAAAAAAAGCTACGACATACGCCAGACATACAGACTACGCTACAGCATACAGCATACGA
CATACGACATACCCCCCAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACACCCCCCCCCCCCCCCCCCCGG
GGGGGGAAAAAGCTACGACATACGCCAGACATACAGACTACGCTACAGCATACAGCATACGACATACGACATACCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCAT
GGGGGGCCCCCCCATAGCATGGGGGGGACGACTACGACATACCATGACTACCAGCATACGACATCATCAGCAGACGACGACGACAGCAGCAGCAGCAGACTTCGACAGCATTTGATA
CGACTACGACGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGCATACGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTTT
TGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAATACATGACGACTGATACTGACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTAC
ACCCCCCGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGCATACGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTTT
GCAAAAGCATAGACTACGACTCAGACGCATAAAAGCTACGACATACGCCAGACATACAGACTACGCTACAGCATACAGCATACGACATACGACATACCCCCCCCCGATGACATACGACA
TACCATAGCATGGGGGGGACGACTACGAAAAAAAAAAGCTACGACATACGCCAGACATACAGACTACGCTACAGCATACAGCATACGACATACGACATACCCCCCCCCGATGACATACG
ACATACCCCCCCCCCCCCATAGCATGGGGCATGACGACTACGACATACAAGTGCGCGGCGCGTTGTGTGTCGCGCGCGCTGGTAACGACAGCGCAGCAGCGCAGACACGATACAGAT
ACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGAC
AGACTACGACAGACTACAGACTACACCCCCCGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGCATACGAACTCAGACGACGACGACTAC
ACAGCGACTAACGACGCATATTTTCAGCACAAAAGGGACGACTACGACATACCATGACTACCAGCATACGACATCATCAGCAGACGACGACGACAGCAGCAGCAGCAGACTTCGACAG
CATTTGATACGACTACGACTACATGACGACTGATACTGAAGACGACTACGACAGACTACGACAGACTACAGACTACACCCCCCCCCCCCCCCCCCCCGGGTTACAGATACGCATACGAC
TACAGACTACGACTACGAATACGACTAAACGGGCATACGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGC
ACAAAAAAACGGGCATACGAACTCAGACGACGACGACTACACAGCGACTAAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGA
CGACTACGACAGACTACGACAGACTACAGACTACACCCCCCCCCCCCCCCCCCCCGGGGGGGGCGCGTGAGATCGTAGATCGTAGGGCGCGTTATTAAGGACGCTGAGGAGCGCTC
GCGTCGAGCGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGCATACGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTT
TGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAGCTACGACACGACGCATATTTTTTTTTTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAGACGACTAC
ACAGCGACTAACGACGCATATTTTTTTTTCTAGCAGATACGCATACAGACTAACGCATACGACATACGCAGCTTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAGACGA
TGATTGACGTAGCATGACGATTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAAAAAAAGCTACGACATACGCCAGACATACAGACTACGCTACAGCATACAGCAT
ACGACATACGACATACCCCCCCCCGATGACATACGACATACCCCCCACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTA
CAGACTACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACACCCCCCCCCCCCCCCCCCCCGGGGGGGGG
GGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGCATACGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTTTTTGAAAAAAAA
AAAGCTACGACACAGACGACTACGACAGACTACGACAGACTACAGACTACACCCCCCCCCCCCCCCCCCCCGATGACATACGACATACCCCCCCCCCCCCATAGCATGGGGGGGACG
ACTACGACATACAAGTACGATACAGATACGACAACAACAATCAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACAAGTACGATACAGATACGACAACAACAAT
CAATGCGATAGACAGACGACTACGACAGACTACGACAGACTACAGACTACACCCCCCGGGGGGGGTTACAGATACGCATACGACTACAGACTACGACTACGAATACGACTAAACGGGC
ATACGAACTCAGACGACGACGACTACACAGCGACTAACGACGCATATTTTTTTTTTTTGCAAAAGCATAGACTACGACTCAGACGCATCAGCACAAAAAAAAAAAAAGCTACGACATCAG
ACGCATCAGCACAAAAAAACGGGCATACGAACTCAGGACTACGACAGACTACAGACTACACC
PAGINA 6 MILIONI
1.000 basi al giorno
1 giorno
tutto il genoma
Quanto siamo diversi ?
Genoma umano: 6.4 x109
paia di basi
the cost to generate a high-quality 'draft' whole human genome sequence
in mid-2015 was just above $4,000; by late in 2015, that figure had
fallen below $1,500. The cost to generate a whole-exome sequence was
generally below $1,000.
I cromosomi umani - cariotipo
E 8-2; 4th 4-10
4th 4-12
Cromosomi
Personalized medicine
takes into account individual genetic differences
• Traditionally, doctors used:
– Family history
– Socioeconomic circumstances
– Environmental factors
• Now:
– genomic/genetic testing
– proteomic profiling
– metabolomic analysis (study metabolites)
Effectiveness of drugs:
Old Paradigm:
New Paradigm:
Medicina personalizzata
Quanto siamo diversi ?
GENOMI
Genoma umano: 6.4 x109 paia di basi
Genomic sequencing
Saccharomyces cerevisiae
[Fungus]
C. elegans
[Worm]
Arabidopsis
[Plant]
Escherichia coli
[Bacterium]
Drosophila
[Insect]
Genomic sequencing
Staph. aureus
[MSRA]
Vibrio cholerae Bacillus anthracis
[cholera]
Yersinia pestis
[Anthrax]
[Plague]
Plasmodium falciparum
[Malaria]
Salmonella
Neisseria meningitidis
[Food poisoning]
[Meningitis]
Cromosomi
Helicobacter pylori
03/10/2005 13.22.00
Assegnato il Premio Nobel per la Medicina 2005, che è andato agli australiani Barry Marshall
e Robin Warren per le loro ricerche sull'Helicobacter pylori. Lo ha reso noto la Nobel Assembly
del Karolinska Institute di Stoccolma. La scoperta dei due ricercatori australiani, avvenuta nel
1982, ha trasformato l'ulcera peptica da infermità cronica a affezione curabile con un breve
regime di antibiotici e altri farmaci, ha valutato il comitato del Premio Nobel. Il premio Nobel per
la Medicina include un assegno di 1,3 milioni di dollari (oltre un milione di euro), un diploma e
una medaglia d'oro. La cerimonia di consegna avverrà in presenza del re di Svezia il 10
dicembre prossimo.
Il genoma umano
Studio del DNA ripetitivo attraverso la
denaturazione e rinaturazione del DNA
Curva di DenaturazioneRinaturazione del DNA eucariotico
Il genoma umano
Tipi di sequenze del genoma
Sequenze ripetute
semplici (Alu)
moderatamente ripetute
trasposoni
retrotrasposoni virali
LINES (long interspersed elements)
SINES (short interspersed elements)
Sequenze ripetute in tandem codificanti per
rRNAs
rRNA 5S
tRNAs
proteine istoniche
Sequenze codificanti per proteine
geni singoli
famiglie geniche
pseudogeni non funzionali
Tipi di sequenze del genoma
Sequenze ripetute
semplici (Alu)
moderatamente ripetute
trasposoni
retrotrasposoni virali
LINES (long interspersed elements)
SINES (short interspersed elements)
Sequenze ripetute in tandem codificanti per
rRNAs
rRNA 5S
tRNAs
proteine istoniche
Sequenze codificanti per proteine
geni singoli
famiglie geniche
pseudogeni non funzionali
DNA moderatamente ripetuto in tandem.
I geni per gli istoni
DNA moderatamente ripetuto in tandem.
I geni per l’RNA ribosomale
La maggior parte del genoma umano è
costituito da sequenze non codificanti
Distribuzione dei geni in diversi organismi
(da genomi compatti a genomi con ampi spazi intergenici)
Struttura dei geni e del genoma umano
Le sequenze
codificanti per
proteine sono una
minima porzione delle
sequenze genomiche
DNA
RNA
5’
3’
5’
3’
AGCT
TCGA
trascrizione
3’
AGCU
traduzione
Proteina
C-ter
N-ter
5’
Un gene una proteina
(le eccezioni confermano la regola)
DNA
gene della -globina
segnale di
poli-adenilazione
Sito di inizio trascrizione
hnRNA
poli-A
Splicing
mRNA
Organization of eukaryotic genes
Regulatory sequence
Exon 1
Intron 1
Exon 2
Intron 2
Exon 3
Intron 3
Organization of genes on a human chromosome
Scale of human genome
If 1bp is 1mm:
The human genome would extend 3200 km.
1 protein coding gene every 300 m.
An average gene would extend for 30 m
(coding sequence: about 1 m)
How the genome is packaged in the nucleus?
Quanto è lungo il nostro DNA ?
Genoma aploide 3.2 x 109 bp
Genoma diploide 6.4 x 109
0,34 nm x 6.4 x 109
???
Se svolgiamo tutto il dna le sue dimensioni lineari sono di circa 2 metri
- cromosomi sono lunghi
circa 1 - 2 cm. Spessore 2 nm
- Il nucleo cellulare è lungo 5
mm
Rapporto con le dimensioni “normali”
Genoma 2 metri x 104
spessore 2 nm x 104
Nucleo 5 mM x 104
20 Km di filamento di DNA
20 mm
5 centimentri
Il DNA del genoma umano è costituito da circa 6x109 COPPIE DI
nucleotidi. Questa lunghissima molecola (metri) è ospitata nel nucleo delle
cellule (mm) in virtù di una raffinato ed organizzato impacchettamento.
Il DNA è associato a proteine e forma la CROMATINA
IL DNA nei cromosomi è
associato a proteine
Gli ISTONI
Istoni
>H2b
mtdkitkkkrnetysiyiykvlrqvhpkigvsskamnimnsfvndlferlvsesynlsnssrskt
ltareiqtsvrlvipgelakhsvsegtkavakyrssi
>H2a
msgrgksgkartkaksrssraglqfpvgrvhrflrkgnyakrvgggapvymaavleyltaeilel
agnaardnkksriiprhlqlavrndeelnkllggvtiaqggvlpniqavllpkktgks
>H3
martkqtarkstggkaprkqlatkaarksapatggvkkphrfrpgtvalreirkyqkstellirk
lpfqrlvreiaqdfktdlrfqssavaalqeaaeaylvglfedtnlcaihakrvtimpkdiqlarr
irgera
>H4
msgrgkggkglgkggakrhrkvlrdniqgitkpairrlarrggvkrisgliyeetrgvlkiflen
virdavtyteharrktvtamdvvyalkrqgrtlygfgg
K = lisina
R=arginina
H= istidina
Nucleosoma:
Ottamero istonico
+
146 bp di dna
1.65 giri
Istone H1
Nucleosoma
H2B
H4
05_23_histone core.jpg
H3
DNA
H2A
“Nucleosome”
• The nucleosome core particle consists of approximately
147 base pairs of DNA wrapped in 1.67 left-handed
superhelical turns around a histone octamer
• Octamer: 2 copies each of the core histones H2A, H2B,
H3, and H4
• Core particles are connected by stretches of "linker DNA",
which can be up to about 80 bp long
ELECTRON MICROGRAPHS
NUCLEOSOMES
INTERPHASE
CHROMATIN
MITOTIC
CHROMATIN
Cromatina e Nucleosoma
cromosoma
metafasico
telaio molecolare
“scaffold”
scaffold
fibre cromatiniche
30 nm
cromatina
“filo di perle”
DNA nudo
Video assemblaggio cromosomi
Epigenetics
• Modern experimental techniques and technologies allow for
the genome-wide study of different types of histone
modifications, shedding light on the role of each one
The histone code
• Example H3K4me3
• H3 is the histone
• K4 is the residue that is modified and its position (K
lysine in position 4 of the sequence)
• me3 is the modification (three-methyl groups
attached to K4)
• If no number at the end like in H3K9ac means only
one group
Different chromatin states
Chromatin structure (and thus, gene expression) depend
also on the post-translational modifications associated with
histones forming nuclesomes