1. Genetica per Scienze Naturali a.a. 03-04 prof S. Presciuttini Inheritance Patterns of Individual Genes (1) 1) MITOSIS We will use the alleles A and a as "typical" alleles of a gene. We can represent gene duplication and segregation as follows: Haploid cells can be of genotype A or a, and the diploids can be homozygous, A/A and a/a, or heterozygous, A/a. Because each of the chromosomes is replicated faithfully, the genotypes of the daughter cells must be identical with the progenitor. Genotype A Genotype a Genotype A/A Genotype A/a Genotipe A/a

2. Genetica per Scienze Naturali a.a. 03-04 prof S. Presciuttini 2) MEIOSIS Meiosis in homozygous meiocytes can produce only one genotype in the four haploid products of meiosis (the tetrad), whereas starting with a meiocyte of genotype A/a, meiosis produces four haploid cells, of which half are A and half are a, as follows: Genotype A/A Genotype A/a Genotipe A/a The reason for this is that in the A/a meiocyte, the A chromosome produces a pair of sister chromatids A/A, and the homologous chromosome produces a pair a/a. These four copies of the gene end up in the four meiotic product cells. This result, was first observed by Mendel. Inheritance Patterns of Individual Genes (2)

3. Genetica per Scienze Naturali a.a. 03-04 prof S. Presciuttini Mendel’s experiments were devised to explain the mechanisms of inheritance: “The object of the experiment was to observe these variations in the case of each pair of differentiating characters, and to deduce the law according to which they appear in successive generations”. Mendel’s experiments were devised to explain the mechanisms of inheritance: “The object of the experiment was to observe these variations in the case of each pair of differentiating characters, and to deduce the law according to which they appear in successive generations”. Mendel's studies constitute an outstanding example of good scientific technique. He chose research material well suited to the study of the problem at hand, designed his experiments carefully, collected large amounts of data, and used mathematical analysis to show that the results were consistent with his explanatory hypothesis. The predictions of the hypothesis were then tested in a new round of experimentation. Mendel's studies constitute an outstanding example of good scientific technique. He chose research material well suited to the study of the problem at hand, designed his experiments carefully, collected large amounts of data, and used mathematical analysis to show that the results were consistent with his explanatory hypothesis. The predictions of the hypothesis were then tested in a new round of experimentation. The techniques of analysis used by Mendel remained unchanged for most part of the XX century; model organisms changed with time, but the basic methodology remained always the same. The existence of genes was originally inferred (and is still inferred today) by observing precise mathematical ratios in the descendants of two genetically different parental individuals. The techniques of analysis used by Mendel remained unchanged for most part of the XX century; model organisms changed with time, but the basic methodology remained always the same. The existence of genes was originally inferred (and is still inferred today) by observing precise mathematical ratios in the descendants of two genetically different parental individuals. Only with the advent of fast DNA sequencing in the 1980’s, genes could be analyzed directly without performing experimental crosses. Only with the advent of fast DNA sequencing in the 1980’s, genes could be analyzed directly without performing experimental crosses. Mendel’s experiments

4. Genetica per Scienze Naturali a.a. 03-04 prof S. Presciuttini Mendel studied the garden pea (Pisum sativum) for two main reasons Mendel studied the garden pea (Pisum sativum) for two main reasons  Peas were available in a wide array of varieties that could be easily identified and analyzed.  Peas can either self (self-pollinate: the male parts - pollen, contained in anthers – can fertilize the female parts – ovules, contained in pistils) or be cross-pollinated (the anthers from one plant are removed before they have opened to shed their pollen, and pollen from the other plant is transferred to the receptive stigma with a paintbrush). Thus, the experimenter can choose to self or to cross the pea plants. Other practical reasons for Mendel's choice of peas were: Other practical reasons for Mendel's choice of peas were:  they are inexpensive and easy to obtain,  take up little space,  have a short generation time,  produce many offspring. Such considerations enter into the choice of organism for any piece of genetic research. Such considerations enter into the choice of organism for any piece of genetic research. The reasons of a choice

5. Genetica per Scienze Naturali a.a. 03-04 prof S. Presciuttini Differences among plant peas The various forms of pea plants available for crossing showed differences in:  length and color of the stem;  size and form of the leaves;  position, color, size of the flowers;  length of the flower stalk;  color, form, and size of the pods;  form and size of the seeds;  color of the seed-coats and of the albumen [cotyledons]. Some of these characters do not permit of a sharp and certain separation, since the difference is of a "more or less" nature, which is often difficult to define. Such characters could not be utilized for the separate experiments; these could only be applied to characters which stand out clearly and definitely in the plants. Lastly, the result must show whether they, in their entirety, observe a regular behavior in their hybrid unions, and whether from these facts any conclusion can be reached regarding those characters which possess a subordinate significance in the type.

6. Genetica per Scienze Naturali a.a. 03-04 prof S. Presciuttini Characters selected for experiments 1.Form of the seeds The depressions, if any, occur on the surface, or they are irregularly angular and deeply wrinkled 2.Color of the seed albumen (endosperm) The albumen is either pale yellow, or it possesses an intense green tint. This difference of color is seen in the seeds as their coats are transparent 3.Color of the seed-coat/ color of the flower This is either white, with which character white flowers are constantly correlated; or it is gray, gray-brown, leather-brown, with or without violet spotting, in which case the color of the standards is violet, that of the wings purple, and the stem in the axils of the leaves is of a reddish tint 4.Form of the ripe pods These are either simply inflated, not contracted in places; or they are deeply constricted between the seeds and more or less wrinkled (P. saccharatum) 5.Color of the unripe pods They are either light to dark green, or vividly yellow, in which coloring the stalks, leaf-veins, and calyx participate 6.Position of the flowers They are either axial, that is, distributed along the main stem; or they are terminal, that is, bunched at the top of the stem and arranged almost in a false umbel; in this case the upper part of the stem is more or less widened in section (P. umbellatum) 7.Length of the stem The length of the stem is very various in some forms; it is, however, a constant character for each, in so far that healthy plants, grown in the same soil, are only subject to unimportant variations in this character. In experiments with this character, in order to be able to discriminate with certainty, the long axis of 6 to 7 ft. was always crossed with the short one of 3/4 ft. to 1 and 1/2 ft

7. Genetica per Scienze Naturali a.a. 03-04 prof S. Presciuttini Plants differing in one character Mendel chose seven different characters to study. The word character in this regard means a specific property of an organism; geneticists use this term as a synonym for characteristic or trait. Mendel chose seven different characters to study. The word character in this regard means a specific property of an organism; geneticists use this term as a synonym for characteristic or trait. For each of the characters that he chose, Mendel obtained lines of plants, which he grew for two years to make sure that they were pure. A pure line is a population that shows no variation in the particular character being studied; that is, all offspring produced by selfing or crossing within the population are identical for this character. For each of the characters that he chose, Mendel obtained lines of plants, which he grew for two years to make sure that they were pure. A pure line is a population that shows no variation in the particular character being studied; that is, all offspring produced by selfing or crossing within the population are identical for this character. These two characters can be directly scored in the crossed plants

8. Genetica per Scienze Naturali a.a. 03-04 prof S. Presciuttini Dichotomous phenotypes Each pair of Mendel's plant lines can be said to show a character difference, i.e. a contrasting difference between two lines of organisms (or between two organisms) in one particular character. Each pair of Mendel's plant lines can be said to show a character difference, i.e. a contrasting difference between two lines of organisms (or between two organisms) in one particular character. Contrasting phenotypes for a particular character are the starting point for any genetic analysis. The differing lines (or individuals) represent different forms that the character may take: they can be called character forms, character variants, or phenotypes. Contrasting phenotypes for a particular character are the starting point for any genetic analysis. The differing lines (or individuals) represent different forms that the character may take: they can be called character forms, character variants, or phenotypes. The term phenotype (derived from Greek) literally means "the form that is shown"; it is the term used by geneticists today. The term phenotype (derived from Greek) literally means "the form that is shown"; it is the term used by geneticists today. The description of characters is somewhat arbitrary. For example, we can state the color-character difference in at least three ways: The description of characters is somewhat arbitrary. For example, we can state the color-character difference in at least three ways:

9. Genetica per Scienze Naturali a.a. 03-04 prof S. Presciuttini Reciprocal crosses In one of his early experiments, Mendel pollinated a purple-flowered plant with pollen from a white-flowered plant. Individuals from which an experiment is started are called the parental generation (P). In one of his early experiments, Mendel pollinated a purple-flowered plant with pollen from a white-flowered plant. Individuals from which an experiment is started are called the parental generation (P). All the plants resulting from this cross had purple flowers. This progeny generation is called the first filial generation (F1 ). The subsequent generations produced by selfing are symbolized F2, F3, and so forth. All the plants resulting from this cross had purple flowers. This progeny generation is called the first filial generation (F1 ). The subsequent generations produced by selfing are symbolized F2, F3, and so forth. Mendel made reciprocal crosses. In most plants, any cross can be made in two ways, depending on which phenotype is used as male or female. Mendel made reciprocal crosses. In most plants, any cross can be made in two ways, depending on which phenotype is used as male or female. For example, the following two crosses are reciprocal crosses:

10. Genetica per Scienze Naturali a.a. 03-04 prof S. Presciuttini Monohybrid crosses Next, Mendel selfed the F 1 plants (the pollen of each flower fertilized its own stigma). He obtained 929 pea seeds from this selfing (the F 2 individuals) and planted them. Next, Mendel selfed the F 1 plants (the pollen of each flower fertilized its own stigma). He obtained 929 pea seeds from this selfing (the F 2 individuals) and planted them. Some of the resulting plants were white flowered; the white phenotype had reappeared. Some of the resulting plants were white flowered; the white phenotype had reappeared. Mendel then did something that, more than anything else, marks the birth of modern genetics: he counted the numbers of plants with each phenotype. Mendel then did something that, more than anything else, marks the birth of modern genetics: he counted the numbers of plants with each phenotype. Mendel counted 705 purple-flowered plants and 224 white-flowered plants. He noted that the ratio of 705:224 is almost exactly a 3:1 ratio (in fact, it is 3.1:1). Mendel counted 705 purple-flowered plants and 224 white-flowered plants. He noted that the ratio of 705:224 is almost exactly a 3:1 ratio (in fact, it is 3.1:1).

11. Genetica per Scienze Naturali a.a. 03-04 prof S. Presciuttini Dominance defined Mendel repeated the experiment for the six other character differences. He found the same 3:1 ratio in the F2 generation for each pair. In all cases, one parental phenotype disappeared in the F1 and reappeared in one-fourth of the F2. Mendel repeated the experiment for the six other character differences. He found the same 3:1 ratio in the F2 generation for each pair. In all cases, one parental phenotype disappeared in the F1 and reappeared in one-fourth of the F2. Mendel used the terms dominant and recessive to describe this phenomenon. The purple phenotype is dominant to the white phenotype and the white phenotype is recessive to purple. Thus the operational definition of dominance is provided by the phenotype of an F1 established by intercrossing two pure lines. The parental phenotype that is expressed in such F1 individuals is by definition the dominant phenotype. Mendel used the terms dominant and recessive to describe this phenomenon. The purple phenotype is dominant to the white phenotype and the white phenotype is recessive to purple. Thus the operational definition of dominance is provided by the phenotype of an F1 established by intercrossing two pure lines. The parental phenotype that is expressed in such F1 individuals is by definition the dominant phenotype.

12. Genetica per Scienze Naturali a.a. 03-04 prof S. Presciuttini A diagram of Mendel’s monohybrid experiments P F1F1 x self 519 sampled and selfed F2F2 F3F3 6022 yellow peas 166 peas with yellow progeny only 2001 green peas green peas with green progeny only 353 peas with 3:1 yellow/green progeny All the F2 greens were evidently pure breeding, like the green parental line; but, of the F2 yellows, two-thirds were like the F1 yellows (producing yellow and green seeds in a 3:1 ratio) and one- third were like the pure- breeding yellow parent. Thus the study of the individual selfings revealed that underlying the 3:1 phenotypic ratio in the F2 generation was a more fundamental 1:2:1 ratio

13. Genetica per Scienze Naturali a.a. 03-04 prof S. Presciuttini Mendel’s theory Mendel's explanation is a classic example of a creative model or hypothesis derived from observation and suitable for testing by further experimentation. He deduced the following explanation: 1. The existence of genes. There are hereditary determinants of a particulate nature. 2. Genes are in pairs. Alternative phenotypes of a character are determined by different forms of a single type of gene. In adult pea plants, each type of gene is present twice in each cell, constituting a gene pair. In different plants, the gene pair can be of the same alleles or of different alleles of that gene. 3. The principle of segregation. The members of the gene pairs segregate (separate) equally into the gametes, or eggs and sperm. 4. Gametic content. Consequently, each gamete carries only one member of each gene pair. 5. Random fertilization. The union of one gamete from each parent to form the first cell (zygote) of a new progeny individual is random; that is, gametes combine without regard to which member of a gene pair is carried.

14. Genetica per Scienze Naturali a.a. 03-04 prof S. Presciuttini The 1:2:1 ratio These points can be illustrated diagrammatically for a general case by using A to represent the allele that determines the dominant phenotype and a to represent the gene for the recessive phenotype (as Mendel did). The use of A and a is similar to the way in which a mathematician uses symbols to represent abstract entities of various kinds. In this figure, these symbols are used to illustrate how the preceding five points explain the 1:2:1 ratio. The members of a gene pair are separated by a slash (/). This slash is used to show us that they are indeed a pair; the slash also serves as a symbolic chromosome to remind us that the gene pair is found at one location on a chromosome pair

15. Genetica per Scienze Naturali a.a. 03-04 prof S. Presciuttini The backcross Mendel's next job was to test his model. He did so in the seed-color crosses by taking an F1 plant that grew from a yellow seed and crossing it with a plant grown from a green seed. Mendel's next job was to test his model. He did so in the seed-color crosses by taking an F1 plant that grew from a yellow seed and crossing it with a plant grown from a green seed.  This type of cross, of an individual of uncertain genotype with a fully recessive homozygote, is now called a testcross or a backcross. Because one of the parents contributes only recessive alleles, the gametes of the unknown individual can be deduced from progeny phenotypes A 1:1 ratio of yellow to green seeds could be predicted in the next generation. If Y stand for the allele that determines the dominant phenotype (yellow seeds) and y stand for the allele that determines the recessive phenotype (green seeds), we can diagram Mendel's predictions, as in the figure beside. A 1:1 ratio of yellow to green seeds could be predicted in the next generation. If Y stand for the allele that determines the dominant phenotype (yellow seeds) and y stand for the allele that determines the recessive phenotype (green seeds), we can diagram Mendel's predictions, as in the figure beside. In this experiment, Mendel obtained 58 yellow (Y /y ) and 52 green (y /y ), a very close approximation to the predicted 1:1 ratio and confirmation of the equal segregation of Y and y in the F1 individual.In this experiment, Mendel obtained 58 yellow (Y /y ) and 52 green (y /y ), a very close approximation to the predicted 1:1 ratio and confirmation of the equal segregation of Y and y in the F1 individual.

16. Genetica per Scienze Naturali a.a. 03-04 prof S. Presciuttini Dihybrid crosses Mendel went on to analyze the descendants of pure lines that differed in two characters. Mendel went on to analyze the descendants of pure lines that differed in two characters. Here we need a general symbolism to represent genotypes including two genes. If two genes are on different chromosomes, the gene pairs are separated by a semicolon, for example, A /a ; B /b. If they are on the same chromosome, the alleles on one chromosome are written adjacently and are separated from those on the other chromosome by a slash, for example, A B /a b or A b /a B. Here we need a general symbolism to represent genotypes including two genes. If two genes are on different chromosomes, the gene pairs are separated by a semicolon, for example, A /a ; B /b. If they are on the same chromosome, the alleles on one chromosome are written adjacently and are separated from those on the other chromosome by a slash, for example, A B /a b or A b /a B. An accepted symbolism does not exist for situations in which it is not known whether the genes are on the same chromosome or on different chromosomes. For this situation, we will separate the genes with a dot, for example, A /a ·B /b. A double heterozygote, A /a · B /b, is also known as a dihybrid. From studying dihybrid crosses (A /a · B /b × A /a · B /b ), Mendel came up with another important principle of heredity. An accepted symbolism does not exist for situations in which it is not known whether the genes are on the same chromosome or on different chromosomes. For this situation, we will separate the genes with a dot, for example, A /a ·B /b. A double heterozygote, A /a · B /b, is also known as a dihybrid. From studying dihybrid crosses (A /a · B /b × A /a · B /b ), Mendel came up with another important principle of heredity.

17. Genetica per Scienze Naturali a.a. 03-04 prof S. Presciuttini Yellow/green-round/wrikled seeds The two specific characters that he began working with were seed shape and seed color. The two specific characters that he began working with were seed shape and seed color. To perform a dihybrid cross, Mendel started with two parental pure lines. One line had yellow, wrinkled seeds; because Mendel had no concept of the chromosomal location of genes, we must use the dot representation to write this genotype as Y /Y · r /r. The other line had green, round seeds, the genotype being y /y · R /R. To perform a dihybrid cross, Mendel started with two parental pure lines. One line had yellow, wrinkled seeds; because Mendel had no concept of the chromosomal location of genes, we must use the dot representation to write this genotype as Y /Y · r /r. The other line had green, round seeds, the genotype being y /y · R /R. The cross between these two lines produced dihybrid F1 seeds of genotype R /r · Y /y, which he discovered were round and yellow. The cross between these two lines produced dihybrid F1 seeds of genotype R /r · Y /y, which he discovered were round and yellow. This result showed that the dominance of R over r and of Y over y was unaffected by the presence of heterozygosity for either gene pair in the R /r · Y /y dihybrid. This result showed that the dominance of R over r and of Y over y was unaffected by the presence of heterozygosity for either gene pair in the R /r · Y /y dihybrid.

18. Genetica per Scienze Naturali a.a. 03-04 prof S. Presciuttini DiHybrid-cross F 2 ratios Next Mendel made the dihybrid cross by selfing the dihybrid F1 to obtain the F 2 generation. The F 2 seeds were of four different types in the following proportions: Next Mendel made the dihybrid cross by selfing the dihybrid F1 to obtain the F 2 generation. The F 2 seeds were of four different types in the following proportions: What could be the explanation? Mendel added up the numbers of individuals in certain F 2 phenotypic classes to determine if the monohybrid 3:1 F2 ratios were still present. He noted that, in regard to seed shape, there were 423 round seeds (315+108) and 133 wrinkled seeds (101+32). This result is close to a 3:1 ratio. Next, in regard to seed color, there were 416 yellow seeds (315+101) and 140 green (108+32), also very close to a 3:1 ratio. The presence of these two 3:1 ratios hidden in the 9:3:3:1 ratio was undoubtedly a source of the insight that Mendel needed to explain the 9:3:3:1 ratio, because he realized that it was nothing more than two independent 3:1 ratios combined at random.

19. Genetica per Scienze Naturali a.a. 03-04 prof S. Presciuttini Visualizing the 9:3:3:1 ratio One way of visualizing the random combination of these two ratios is with a branch diagram, as follows: One way of visualizing the random combination of these two ratios is with a branch diagram, as follows: The combined proportions are calculated by multiplying along the branches in the diagram because, for example, 3/4 of 3/4 is calculated as 3/4 × 3/4, which equals 9/16 These multiplications give us the following four proportions: The combined proportions are calculated by multiplying along the branches in the diagram because, for example, 3/4 of 3/4 is calculated as 3/4 × 3/4, which equals 9/16 These multiplications give us the following four proportions:

20. Genetica per Scienze Naturali a.a. 03-04 prof S. Presciuttini The Punnett square The four female gametic types will be fertilized randomly by the four male gametic types to obtain the F2, and the best way of showing this graphically is to use a 4×4 grid called a Punnett square, which is depicted in Figure 2-10. Grids are useful in genetics because their proportions can be drawn according to genetic proportions or ratios being considered, and thereby a visual data representation is obtained. In the Punnett square in Figure 2-10, for example, we see that the areas of the 16 boxes representing the various gametic fusions are each one-sixteenth of the total area of the grid, simply because the rows and columns were drawn to correspond to the gametic proportions of each. As the Punnett square shows, the F2 contains a variety of genotypes, but there are only four phenotypes and their proportions are in the 9:3:3:1 ratio. The four female gametic types will be fertilized randomly by the four male gametic types to obtain the F2, and the best way of showing this graphically is to use a 4×4 grid called a Punnett square, which is depicted in Figure 2-10. Grids are useful in genetics because their proportions can be drawn according to genetic proportions or ratios being considered, and thereby a visual data representation is obtained. In the Punnett square in Figure 2-10, for example, we see that the areas of the 16 boxes representing the various gametic fusions are each one-sixteenth of the total area of the grid, simply because the rows and columns were drawn to correspond to the gametic proportions of each. As the Punnett square shows, the F2 contains a variety of genotypes, but there are only four phenotypes and their proportions are in the 9:3:3:1 ratio.

21. Genetica per Scienze Naturali a.a. 03-04 prof S. Presciuttini Chi-square analysis of Mendel’s single-plant observations Mendel reported the following results for the F 2 progeny of single plants: