Conducted pea research between 1856 and 1863 In 1866 he published Experiments in Plant Hybridization, (Versuche über Pflanzen- Hybriden) in which he

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1 Gregor Johann Mendel Austrian Monk, born in what is now Czech Republic in 1822 Son of peasant farmer, studied Theology and was ordained priest Order St. Augustine. Went to the university of Vienna, where he studied botany and learned the Scientific Method Worked with pure lines of peas for eight years Pea experiments had been conducted centuries earlier in England, but were poorly interpreted

2 Conducted pea research between 1856 and 1863 In 1866 he published Experiments in Plant Hybridization, (Versuche über Pflanzen- Hybriden) in which he established his three Principles of Inheritance Work was largely ignored for 34 years, until 1900, when 3 independent botanists rediscovered Mendel s work. (De Vries, von Tschermak & Correns)

3 Mendel was the first biologist to use mathematics to explain his results quantitatively. Mendel predicted The concept of genes That genes occur in pairs That one gene of each pair is present in the gametes

4 Genetics terms you need to know: Gene a unit of heredity; a section of DNA sequence encoding a single protein Genome the entire set of genes in an organism Alleles two genes that occupy the same position on homologous chromosomes and that cover the same trait (like flavors of a trait). Locus a fixed location on a strand of DNA where a gene or one of its alleles is located.

5 Homozygous having identical genes (one from each parent) for a particular characteristic. Heterozygous having two different genes for a particular characteristic. Dominant the allele of a gene that masks or suppresses the expression of an alternate allele; the trait appears in the heterozygous condition. Recessive an allele that is masked by a dominant allele; does not appear in the heterozygous condition, only in homozygous.

6 Genotype the genetic makeup of an organisms Phenotype the physical appearance of an organism (Genotype + environment) Monohybrid cross: a genetic cross involving a single pair of genes (one trait); parents differ by a single trait. P = Parental generation F 1 = First filial generation; offspring from a genetic cross. F 2 = Second filial generation of a genetic cross

7 Mendel s Principles 1. Principle of Dominance: One allele masked another, one allele was dominant over the other in the F 1 generation. 2. Principle of Segregation: When gametes are formed, the pairs of hereditary factors (genes) become separated, so that each sex cell (egg/sperm) receives only one kind of gene.

8 Principle of Independent Assortment Based on the pea results, Mendel postulated the 3. Principle of Independent Assortment: Members of one gene pair segregate independently from other gene pairs during gamete formation Genes get shuffled these many combinations are one of the advantages of sexual reproduction

9 A Warning on Assortment Today we know independent assortment works only if the genes lie on different chromosomes If two genes lie on the same chromosome, they will be transmitted together Mendel looked at seven traits he reported as independently assorted. His peas had seven pairs of chromosomes. Historians say he likely threw away data that did not fit his hypotheses!

10 Coming Next: Incomplete dominance Human blood types Complex traits Sex-linked traits

11 Dominanza incompleta o codominanza Quando nell eterozigote i due alleli si esprimono entrambi in egual misura e l espressione di ciascun allele è riconoscibile a livello fenotipico. Esempio: gruppi sanguigni sistema ABO Genotipo I A I A I A i I B I B I B i I A I B i - i Fenotipo Gruppo A Gruppo B Gruppo AB Gruppo 0

12 Penetranza: la probabilità che un allele si esprima negli individui che lo possiedono; Penetranza completa: se il 100% degli individui portanti un determinato allele manifestano il fenotipo corrispondente; Penetranza ridotta o incompleta: se la frequenza di espressione è inferiore al 100%. Espressività: grado di manifestazione del carattere; Espressività uniforme: il carattere fenotipico è uguale in tutti gli individui Espressività variabile: manifestazione fenotipica differente in individui con lo stesso genotipo; :

13 Geni localizzati sul cromosoma X La trasmissione ereditaria dei geni X-linked è diversa da quella dei geni autosomici in quanto si osserva differenza tra gli incroci reciproci, cioè la F1 è diversa a seconda che un carattere sia trasmesso dal padre o dalla madre.

14 Punnett Square for Sex Determination Reginald Punnett ( ) developed this device to explain sex determination. He explored sex-linked coloration in chickens. Female gametes across top Male gametes along side

15 Inattivazione del cromosoma X nelle cellule di mammifero

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18 Modelli di Ereditarietà Autosomica Il gene la cui mutazione è responsabile dell insorgenza del fenotipo è localizzato sugli autosomi X-linked Il gene è localizzato sul cromosoma X (differente l espressione nei due sessi) Y-linked Il gene è localizzato sul cromosoma Y (Eredità paterna) Mitocodriale Il fenotipo è determinato da geni localizzati nel genoma mitocondriale (Eredità materna)

19 Modelli di Ereditarietà Dominante L eterozigote manifesta il fenotipo (guadagno di funzione) Recessivo Soltanto l omozigote manifesta il fenotipo (perdita di funzione)

20 Autosomica Dominante Neurofibromatosi Corea di Huntington Autosomica Recessiva Talassemie Falcemia Fibrosi cistica Fenilchetonuria X-linked Distrofia muscolare Cecità ai colori Favismo X-fragile

21 Year Disease MIM n Location Gene Chromosome abnormality 1986 Duchenne muscular dystrophy Xp21.3 DMD (a) del(x)(p21.3) (b) t(x;21)(p21.3:p13) Retinoblastoma q14 RB del(13)(q13.1q14.5) 1989 Cystic fibrosis q31 CFTR None 1990 Neurofibromatosis q11.2 NF1 Balanced translocations t(1;17)(p34.3:q11.2) t(17;22)(q11.2:q11.2) Wilms' tumor p13 WT1 del(11)(p14p13) 1991 Aniridia p13 PAX6 t(4;11)(q22;p13) del(11)(p13) Familial polyposis coli q21 APC del(5)(q15q22) Fragile-X syndrome Xq27.3 FMR1 FRAXA fragile site Myotonic dystrophy q13.3 DMPK None 1993 Huntington's disease p16 HD None Tuberous sclerosis p13 TSC2 Microdeletions in candidate region von Hippel-Lindau disease p25 VHL Microdeletions in candidate region 1994 Achondroplasia p16 FGFR3 None Early-onset breast/ovarian q21 BRCA1 None cancer Polycystic kidney disease p13.3 PKD1 t(16;22) (p13.3;q11.21) Spinal muscular atrophy q13 SMN1 None

22 OMIM Statistics for May 26, 2008 Autosomal X-Linked Y-Linked Mitochondrial Total * Gene with known sequence Gene with known sequence and phenotype # Phenotype description, molecular basis known % Mendelian phenoty pe or locus, molecular basis unknown Other, mainly phenotypes with suspected mendelian basis Total

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25 (A) Autosomal dominant; (B) autosomal recessive; (C) X-linked recessive; (D) X-linked dominant; (E) Y-linked.

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32 Genoma Mitocondriale bp 37 geni

33 Eredità mitondriale o Eredità materna LEBER HEREDITARY OPTIC NEUROPATHY; LHON

34 Human case: CF Mendel s Principles of Heredity apply universally to all organisms. Cystic Fibrosis: a lethal genetic disease affecting Caucasians. Caused by mutant recessive gene carried by 1 in 20 people of European descent (12M) One in 400 Caucasian couples will be both carriers of CF 1 in 4 children will have it. CF disease affects transport in tissues mucus is accumulated in lungs, causing infections.

35 Inheritance pattern of CF IF two parents carry the recessive gene of Cystic Fibrosis (c), that is, they are heterozygous (C c), one in four of their children is expected to be homozygous for cf and have the disease: C c C C = normal C c = carrier, no symptoms c c = has cystic fibrosis C c C C C c C c c c

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39 Neuropsychiatric diseases caused by expansion of trinucleotide repeats Myotonic dystrophy Fragile X syndrome Spinal and bulbar muscular atrophy (Kennedy s) Huntington s disease Microsatellites short regions of repeating DNA sequence in the genome (because their G+C content is usually higher or lower than the average for the genome they frequently appear to band at a different buoyant density in CsCl gradients and hence are called satellites ) microsatellites are often comprised of trinucleotide repeats

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41 Frequenza: 1/4000 maschi. X fragile (309550) Ereditarietà: Legata al cromosoma X. Malattia causata da mutazione dinamica. Genetica: Nel 1991 è stato identificato il gene responsabile. La mutazione è caratterizzata dall amplificazione di un tratto di DNA costituito da una specifica sequenza ripetuta (CGG). Nei soggetti normali è presente un numero di ripetizioni variabili da 6 a 55. Esistono due differenti tipi di mutazione: la premutazione (56-200) e la mutazione completa (>200). La probabilità di espansione aumenta con le dimensioni della premutazione e quindi con il passare delle generazioni (Paradosso di Sherman). Diagnosi: La diagnosi molecolare (Southern blot) permette di individuare anche gli individui con la premutazione.

42 Malattia di Huntington (143100) Frequenza: 5-10/ nati vivi Ereditarietà: autosomica dominante. Malattia causata da mutazione dinamica Genetica: Il gene responsabile della malattia ed il suo prodotto proteico sono stati identificati. Il gene definito Intersting Transcript (IT-15), è localizzato sul braccio corto del cromosoma 4 (4p16.3). La malattia è associata all amplificazione patologica di una specifica sequenza ripetuta (CAG) nell allele mutato. Nella popolazione normale la tripletta è ripetuta volte. Nei pazienti affetti il numero di ripetizioni varia da 36 a più di 100. Un numero intermedio di espansioni volte, è considerato una premutazione. Diagnosi: Il test genetico si basa sulla determinazione del numero di espansione della tripletta.

43 Human Genome Project February 2001 Studio delle malattie genetiche

44 Classificazione????????????

45 Numero di geni Malattie monogeniche e Fenotipi complessi Tipo cellulare Zigote o mutazioni somatiche Funzione del gene interessato

46 Malattie congenite La mutazione è presente nello zigote Malattie ad esordio tardivo La mutazione (o le mutazioni) insorge durante la vita dell individuo in cellule somatiche

47 Mutazioni somatiche e germinali

48 Malattie congenite Mutazioni geniche Alterazioni complesse del genoma (mutazioni cromosomiche)

49 Malattie ad esordio tardivo Cancro Malattie degenerative Malattie infettive

50 Presenti nello zigote Alterazioni complesse del genoma Mutazioni cromosomiche

51 This database is a catalog of human genes and genetic disorders authored and edited by Dr. Victor A. McKusick and his colleagues at Johns Hopkins and elsewhere, and developed for the World Wide Web by NCBI, the National Center for Biotechnology Information. The database contains textual information and references. It also contains copious links to MEDLINE and sequence records in the Entrez system, and links to additional related resources at NCBI and elsewhere.

52 Studio delle malattie genetiche Dal Fenotipo al Genotipo Dal Genotipo al Fenotipo

53 Studio delle malattie genetiche Dal Fenotipo al Genotipo Dal Genotipo al Fenotipo

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56 Commonly used methods for identifying genes in cloned DNA Zoo blotting Method Comments A DNA clone is hybridized at reduced hybridization stringency against a Southern blot of genomic DNA samples from a variety of animal species, a zoo blot. Depends on coding DNA being more strongly conserved in evolution than non-coding DNA (Figure 10.21). Many vertebrate genes have associated CpG islands, hypomethylated GCrich sequences usually having multiple rare-cutter restriction sites ( Cross and Bird, 1995). CpG island identification Identification by restriction mapping. DNA clones are usually hybridized against Southern blots of genomic DNA cut with SacII, EagI or BssHII to identify clustering of rare-cutter sites (Figure 10.22). Island-rescue PCR. This is a way of isolating CpG island sequences from YACs by amplifying sequences between islands and neighbouring Alu repeats. Hybridization Exon trapping cdna selection or capture Computer analysis of DNA sequence A genomic DNA clone can be hybridized against a Northern to mrna/cdna blot of mrna from a panel of culture cell lines, or against appropriate cdna libraries. This is essentially an artificial RNA splicing assay (see Figure 10.23). It relies on the observation that the vast majority of mammalian genes contain multiple exons which need to be spliced together at the RNA level. These techniques involve repeated purification of a subset of genomic DNA clones which hybridize to a given cdna population (see Figure 10.24). Homology searches. Any DNA sequence obtained from a genomic clone can be compared against all other sequences in sequence data-bases. Significant homology to known coding DNA or gene-associated sequences may indicate a gene (see Section ) Gene searching algorithms. A variety of computer programs have been developed to search sequences for exons and other gene-associated motifs (see Figure and Section ).

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58 Year Disease MIM n Location Gene Chromosome abnormality 1986 Duchenne muscular dystrophy Xp21.3 DMD (a) del(x)(p21.3) (b) t(x;21)(p21.3:p13) Retinoblastoma q14 RB del(13)(q13.1q14.5) 1989 Cystic fibrosis q31 CFTR None 1990 Neurofibromatosis q11.2 NF1 Balanced translocations t(1;17)(p34.3:q11.2) t(17;22)(q11.2:q11.2) Wilms' tumor p13 WT1 del(11)(p14p13) 1991 Aniridia p13 PAX6 t(4;11)(q22;p13) del(11)(p13) Familial polyposis coli q21 APC del(5)(q15q22) Fragile-X syndrome Xq27.3 FMR1 FRAXA fragile site Myotonic dystrophy q13.3 DMPK None 1993 Huntington's disease p16 HD None Tuberous sclerosis p13 TSC2 Microdeletions in candidate region von Hippel-Lindau disease p25 VHL Microdeletions in candidate region 1994 Achondroplasia p16 FGFR3 None Early-onset breast/ovarian q21 BRCA1 None cancer Polycystic kidney disease p13.3 PKD1 t(16;22) (p13.3;q11.21) Spinal muscular atrophy q13 SMN1 None

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61 Human Genome Project Studio delle malattie genetiche Diagnosi Cura Prevenzione

62 Multiplex ARMS test per individuare 4 specifiche mutazioni che causano la fibrosi cistica

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64 Diagnosi di malattie da espansione di triplette ripetute

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70 MEDICINA MOLECOLARE