1

BASICS OF GENETICS
1.1 PRELIMINARIES

It is a matter of general observation that the cubs of a lion resemble the lion and pups of a dog resemble the dog not only in shape but also in habits, strength and functional abilities. This phenomenon was described by a phrase like begets like or chips of the same block. Evidently, there is something common between the parents and their offspring which was responsible for such a tidy resemblance. Many speculations were made but none turned out to be correct unless mode of heredity in sexually reproducing animals was unraveled. It was Johann Gregor Mendel who gave the concept of factors being transmitted from parents to offsprings. These factors later turned out to be the structural units called genes. Interestingly, variation is also a part of heredity because no two individual are identical in every respect. Even homozygous twins may differ in size, weight and in appearance due to acquired characters. Thus, similarity and variation go hand in hand as the law of nature and constitute the raw material for a geneticist and breeder to work upon and evolve new strains, breeds, varieties etc. It is not very far when gene was a hypothetical unit lying on the chromosome and somehow governing the expression of traits in an individual and imparting immortality to an individual by keeping alive its characters from generation after generation albeit in different proportions. Breeding experiments were the only tool available to a geneticist to speculate the genetic constitution of an individual and plan his experiments for better production. It is only half of the century when the chemical nature of the gene was explored and they can now be replicated in a test tube, their nature can be altered and they can be commanded to perform a desired function. In spite of these epoch making discoveries, the fundamentals of genetics as laid down by early scientists have not changed and even today constitute the basis of green revolution in agriculture and white revolution in milk production. The purpose of this chapter is not to elaborate the theoretical part of genetics but to familiarise the readers with the basics of genetics and various terminologies used in the sequel. For in-depth knowledge of the principles of genetics, the readers are advised to refer the books given at the end of this chapter. The universe consists of nonliving objects (inanimates) and living organisms (animates). The former are static, lifeless, nonsensual and nonreproducing whereas the latter have life, sensation, dynamism and are endowed with the capacity to reproduce. The structural and functional unit of all living organisms is the cell. The cell theory was propounded by Schleiden and Schwann in 1838 and Virchow in 1855 which stated that all organisms are composed of one or more cells and these cells arise only from preexisting cells. An organism may be as small as unicellular or as big as an elephant consisting of billions of trillion cells.
1

STATISTICAL ANALYSIS OF QUANTITATIVE GENETICS

Cell
It is a microscopic unit circumscribed by a cell membrane in animals or a true cell wall in plants. The inner material (protoplasm) consists of a dense mass in the centre called nucleus and a jelly like thick matrix surrounding the nucleus called cytoplasm. The latter contains many submicroscopic organelles like mitochondria, golgi bodies, centrosome, lysosomes, endoplasmic reticulum etc., which perform specialized functions in the cell. The nucleus contains a sap the nucleoplasm and thread like bodies termed as chromosomes. The chromosomes are the structural unit of inheritance and carry many genes on them which are the functional unit of a trait or character. The location on the chromosome where a gene rests is termed as locus. Each species has a specific number of chromosomes ranging from 4 in drosophila fly to 254 in shrimps (Eupagurus ochotensis). In human beings, the number is 46 (23 pairs). Chromosomes always occur in pairs. All cells of the body barring germ cells are called somatic cells and are characterised by the presence of the number of chromosomes specific for the species. On the other hand, gametic cells (ova, produced by ovary in female and sperm, produced by testes in male) contain half the number of chromosomes present in a somatic cell deriving one member from each pair of chromosomes. In numerical terms, if n represents the number of pairs of chromosomes, then a somatic cell will have 2n chromosomes and the condition is known as diploid number whereas a gametic cell has only n chromosomes and is termed as haploid number. Occasionally, more than two chromosomes are present in a pair and depending on its number, i.e., 3 or 4, the condition is known as triploid or tetraploid or in a general term polyploid. Broadly, the chromosomes have been classified as autosomes ( n 1)pairs) and sex chromosomes (one pair). The former determine the traits of an individual while the latter are responsible for determining the sex of the individual. Again the sex chromosomes are of two types, X type and Y type. The presence of two X (XX) chromosomes gives birth to a female, the condition is designated as homogametic. Thus, the female produces only one type (X type) of gametes. On the other hand a XY composition symbolises a male, the condition is called heterogametic and the individual produces two types of gametes viz., X type and Y type. Thus, all females are homogametic and all males are heterogametic. A condition of YY chromosomes does not exist. Rarely abnormal individuals such as XXY (Klinfelter s syndrome) and XO (Turner s syndrome) have been detected. They are always sterile.

G ene
Genes are the structural and functional units lying on the chromosomes. Like chromosomes they exist in pairs, one on each member of the chromosome and determine the biological character of an individual. The two sister genes on a particular locus may or may not be identical but are always complementary to each other. They are technically known as alleles, the abbreviated form of the word allelomorph. Chemically, the genes are composed of deoxyribonucleic acid (DNA) and direct the cell to perform a particular function by using chemical triplet code by way of messenger ribonucleic acid (mRNA). There are a large number of genes existing on each chromosome but their exact number is not known. According to a rough estimate, by the use of chromosomal maps, the number of genes in human beings has been found to be around 30,000 which may be far from reality. Bo-yuan and his colleagues of Ohio State University estimated that the Human Genome Consortium is 65,000 75,000.

BASICS OF GENETICS

Cell Division
There are two chief processes of cell division (i) Mitosis (ii) Meiosis.

Mitosis
In the process of mitosis which is normally present in all somatic cells, the sister chromosomes of each pair become distinct, thickened and arrange themselves in a single plane about the centre of the cell. Each chromosome is then duplicated, when for a short time the number of chromosomes is doubled with their genes. One of the replicated chromosome passes to each pole of the cell. The cell wall constricts and the cytoplasm is divided into two equal halves giving rise to two daughter cells which are exactly similar to that of the mother cell and each daughter cell carries diploid number of chromosomes. Thus, mitosis is the process of increasing the cell number without disturbing the genetic constitution of the cell.

M eiosis
This is also known as the reduction division. The main feature of this process of cell division is that one partner of each pair of chromosomes passes into daughter cells with the result that each daughter cell has half the number of chromosomes (n) of the parent cell. This process is observed in gametogenic cells and the gametes (ova, sperm) so formed have only half the number of chromosomes (haploid number).

Reproduction
As mentioned earlier, the living organisms possess the capacity to reproduce i.e., to create the new ones of their own kind to maintain the continuity of the species in space and time. Reproduction is also a necessity to replace the individuals which grow old and die. In general, two types of reproduction has been observed namely, (i) asexual (ii) sexual.

Asexual Reproduction
This type of reproduction is normally observed in unicellular organisms and certain plants. Here an organism is divided into two parts such that each part is regenerated into complete organism similar to that of the parent organism. In some plants, a small portion such as root, stem or leaf may establish itself as a new plant. This process does not contribute to genetic variability and hardly holds any importance in genetic analysis.

Sexual Reproduction
This occurs in animals, human beings and plants as well as in organisms where distinct dimorphism exists i.e., where male and female individuals can be distinctly identified. It can be said that the vignettes of reproduction would have not been so prismatic, had there been no sexual reproduction. In bisexual animals, reproduction is possible only with conjugal cooperation and coordination of the female and male. The male gonad, the testis produces male gametes called sperms. The peculiarities of the male gametes are : ( i ) they are produced in large numbers (in billions). (ii ) they are unicellular in nature.

STATISTICAL ANALYSIS OF QUANTITATIVE GENETICS

(iii) they are motile. ( iv ) each gamete contains half of the number of chromosomes. ( v ) they are heterogametic i.e., half of the sperms produced contain X chromosomes and the other half Y chromosomes. On the other hand, the female gamete, Ovum is produced by the ovary and it contains the following properties. ( i ) only one or few ova are produced at a time. (ii ) each ovum has half the number of chromosomes (n) and they are homogametic i.e., all the ova produced contain X-chromosomes. (iii) ovum is unicellular. ( iv ) ovum is nonmotile. After pairing, the male and female gametes unite restoring the normal 2n number of chromosomes. If the ovum with X chromosome happens to unite with the sperm containing X chromosome, the offspring produced is a female. But if the ovum unites with a sperm having Y chromosome, the resulting young one is a male. Thus, the bridge between the offspring and the parents are the chromosomes with their genes which determine the traits or characters of an individual. Hence, the phenomenon of transmission of traits from parents to offsprings is known as heredity and the science dealing with the resemblance and differences among the related organisms has been termed as Genetics (Beatson 1905).

1.2

FUNDAMENTALS OF GENETICS

It is a matter of common experience that the individuals of different species differ widely both in structure and function. Not only that, even the individuals within a species are highly variable. The offsprings inherit characters from their parents but still neither the parents and their offsprings, nor the real brothers and sisters are identical in all respects. The variation between individuals is attributed compositely to genetic composition, environmental factors and their interaction. A number of theories were propounded from time to time to explain the possible mechanism of inheritance based on experience and speculations, but none stood the test of time. Later, the theory of quantitative or biometrical genetics came into existence on the basis of experiments conducted by Gregor J. Mendel honoured as the Father of Genetics on garden pea in 1866. Mendel proposed two laws of inheritance which were not recognised until the same were rediscovered and reaffirmed by Derries in Holland, Correns in Germany and Tschermark in Austria in 1900. Mendel died in 1884 and never lived to see that he had opened the water gates to a new science called genetics. Mendels first principle of inheritance is called the law of segregation. It states that the allelic genes in zygote do not blend or contaminate each other but segregate and pass into different gametes. The segregation occurs at the meotic division. Mendels second principle is known as the law of independent assortment. It states that, during meiosis one of the chromosome in the pair is contributed independently in the gamete without being influenced by other chromosomes or cytoplasmic factors. To explain this law, it may be recalled that the chromosomes and the genes always exist in pairs (all allelomorph) and at the time of reduction division during gamete formation, any one partner of the pair goes to one gamete and the other to the other gamete. Thus, the gametes formed may have similar number of chromosomes

BASICS OF GENETICS

and the genes but may not be identical. To elaborate it further, suppose a species has two pairs of chromosomes with one allelic gene on each pair of chromosome as follows :

Thus, if there are two pairs of allelic genes, four types of gametes with different genetic composition are formed, provided both the pairs are heterozygous. If the number of allelic pairs is n, the possible number of gametes is 2n. This is due to uninhibited combination of any one allele for each pair of gene and is called independent assortment. A third principle recognised by Mendel is called law of dominance. The law emphasises that the two alleles of a particular gene may not be identical and may impart different character to a particular trait e.g., one allele of a particular gene, say R, may impart red colour to a flower while its allele, r, may be responsible for white colour to the flower. When either is present in a homozygous condition, the colour of the flower is either red (RR) or white (rr). On the other hand, if they are present in a heterozygous condition (Rr), the expression of the white colour is masked by the dominant red colour and the flower emerges as red. In this case, red colour allele (R) is said to be dominant and white colour gene (r) is said to be recessive. From the above, it is evident that an organism inherits genes from its parents which are responsible for the expression of characters. However, these genes are not expressed in vacuum but need proper environment to manifest their maximal capacity. If the required environment is not available, the expression of the character may be inhibited fully or partially. For example, a plant may have the gene to grow tall but if the soil is deficient in nitrogen, minerals and water, the plants growth may remain stunted belying the presence of gene for tallness. Thus, an individual can be deemed as genotype depending on the genetic material inherited from its parents and what it appears to be is phenotype. Ideally, the genotype can be defined as the inherited genetic constitution or the sum total of heredity that an individual receives from its parents. Thus the total genic composition (genome) carries the blue print of structure and function for an individual. On the other hand, a phenotype is the appearance or performance of an organism as a result of the interaction of a genotype with that of a given environment. In nut shell, a genotype determines, what an individual should be while phenotype reveals what it is. Further, a genotype is constant from birth till death whereas phenotype changes with time, place, age, food, environment etc.

Qualitative and Quantitative Characters

With the gain in knowledge about genes, their interaction and expression, it may be emphasized that all characters are not controlled by a single gene but a trait may be controlled by one, two, three or more genes. Accordingly, the traits may be classified into (i) qualitative characters (ii) quantitative characters. Qualitative characters are those which are characterised by the presence or absence of a trait. They are governed by one, two or at the most three pairs of genes. Examples of such characters are, presence or absence of flowers of a particular colour, shape of comb in male fowl, plumage colour in Andalusian fowl, shape of seeds etc. Quantitative characters, on the other hand, are characterised by the presence of continuous type of variation (range) and are controlled by multiple number of genes, each gene contributing its

STATISTICAL ANALYSIS OF QUANTITATIVE GENETICS

share to the totality of the character. Mendelian laws can not directly be applied to such characters. Examples of quantitative characters are milk yield of a cow, yield of a crop, egg production, size of fruits, etc.

Qualitative Genetics
It deals with the qualitative characters. The individuals are classified into a number of distinct classes and as such the qualitative variation can be studied by considering the frequencies into various classes. To test whether the observed frequencies are in agreement with the hypothetical frequencies based on Mendelian laws, one usually applies chi-square test.

Quantitative Genetics
It deals with the quantitative characters of a population. Inferences are drawn about the population from samples collected from the same population. Each individual is represented by a numerical value usually considered as a continuous variable and all statistical tools used for measuring variation of continuous variable(s) can be applied. Normally, the methods involved are mean, variance, analysis of variance, correlation and regression, path coefficients, etc.

Test Crossings
Crossing between individuals of prebreeding history or known genetic constitution for a particular trait is known as the test crossing. This is a useful tool to explore the ingenuity of the genic nature of a character. Depending on the number of genes involved in governing a trait, different terminologies have been used. A cross between two individuals controlled by a single pair of genes is called a monohybrid cross. In this case, if both male and female are homozygous dominant, all offsprings will be homozygous dominant as depicted in Fig. (1.2.1).

Fig. 1.2.1 All homozygous tall.

Similarly, if both sexes are homozygous recessive, all offsprings will be true homozygous recessive. The same phenomenon is exhibited in Fig. (1.2.2).

Fig. 1.2.2 All homozygous recessive.

On the other hand, if one mate is homozygous dominant and the other is homozygous recessive, all offsprings will genetically be heterozygous but will reveal dominant character because the presence of dominant gene masks the expression of recessive gene as presented diagrammatically in Fig. (1.2.3).

BASICS OF GENETICS

Fig. 1.2.3 All heterozygous tall.

But the selfing among F1 generation to yield F2 generation results into a genotypic ratio of 1 : 2 : 1 and phenotypic ratio 3 : 1. The proportion between different types of individuals in F2 generation is known as monohybrid ratio. A cross between F1 generation is shown in Fig. (1.2.4).

Fig. 1.2.4 Homozygous tall

Heterozygous tall

Homozygous recessive

Another method to test the genotype for a monohybrid cross is to cross an F1 individual with its homozygous recessive parent (tt). This is known as back cross or test cross. As a special case, two crosses, A B in which A has the female parent of P1 and male parent of P2 and vice-versa are known as reciprocal crosses. They are of great utility in understanding the genetic composition of monohybrid characters. The crosses involving two pairs of genes or two characters differentiably separable in inheritance is called a dihybrid cross. Similarly, a cross involving three pairs of genes or three characters is known as trihybrid cross and those involving many genes are called polyhybrid crosses.

Genetic and Phenotypic V ariation Variation

Variation is the law of nature. All living beings whether plants, animals or humans differ in shape, size, colour, appearance and functions from each other. This difference is called variation and can be classified into genetic and phenotypic variation.

Genetic V ariation Variation

It is well documented that each character of an individual is governed by one or more number of genes. Further, it has been established that species has particular number of chromosomes with thousands of genes on them. The genetic orientation of these genes varies from individual to individual and gets multiplied in each generation as per the laws of segregation and independent assortment advocated by Mendel. Another factor which constitutes to genetic variation is the sudden alteration in genic structure due to natural or man made causes such as X-rays, ultra violet rays, radiations from radioisopic substances such as radium, uranium, etc. These genic modifications are called mutations. Changes caused by mutations are permanent and heritable to next generation. Thus, the variation resulting due to difference in genetic constitution between individuals is called genetic variation. As explained later, the genetic variation provides raw material for selection to improve the variety of a trait in plants or animals.

STATISTICAL ANALYSIS OF QUANTITATIVE GENETICS

Phenotypic V ariation Variation

As discussed, genes are the basic architects responsible for the characters of an organism but they do not express in vacuum. They need proper environment to manifest themselves. The gene expression becomes modified depending on the suitability of the environment to which an organism is exposed. Thus, the variation brought about by the environment effect(s) is called phenotypic variation. This fact can be demonstrated by the illustration that two cuttings taken from a chrysanthemum plant will have exactly the same genetic constitution. Let these be planted in two separate pots. If one is provided with adequate manure and water, while the other is deprived of these essentials, it will be observed that the former grows rapidly whereas the later has poor or no growth. This substantiates the role of environment in gene expression. It is emphasized that the genotypic variation is inherited but phenotypic variation is temporary and is not transmitted to the next generation. Thus, in the sequel it will be discussed that how the total variation observed between individuals can be attributed and partitioned into genetic variation, phenotypic variation and the variation due to interaction between genetic and environmental effects. Further, how can it be harnessed to improve the quality of a trait by using selective breeding as a tool.

1.3

PROBABILITY AND HEREDITY

Laws of probability are of great help to determine the chance of getting an offspring having certain desired traits. To delve deep on probability, an inquisitive mind should read a book on probability theory namely, Feller, W. (1966), Haight, F.A. (1981). One is often anxious to know the sex of a foetus, the colour of a flower, shape of corn, etc. From the angle of probability, it will not be wrong to assume that the probability of a male birth and that of a female is half and half. We consider the case where a couple wants to know, what is the chance that out of two births, both the children will be boys. If first birth is a male, the sex of the child at second birth is independent of the sex of the child at first birth. So the probability of both 1 1 1 the children being male, by multiplicative law of probability, is = . Again the probability of 2 2 4 1 1 a boy and a girl is . Probability is obtained by the consideration that the probability of a boy 2 2 1 1 1 at first birth and of a girl at second birth is = and that of a girl at first birth and of a boy 2 2 4 1 1 1 1 1 1 at second birth is also = . Hence, the probability of a boy and a girl is + = . Similarly, 2 2 4 4 4 2 the probabilities in case of three births to a couple having all the boys, two boy and one girl, one boy and two girls or all the three girls can be calculated by the respective terms of the binomial expansion of (p + q)3 where p is the probability of a male birth and q of a female birth, each equal 1 to . We know, (p + q)3 = p3 + 3p2q + 3pq2 + q3. On substituting the value of p and q, the 2 1 3 3 1 probabilities of the four events are, , , and respectively. 8 8 8 8

BASICS OF GENETICS

Consider another problem. In a couple, the wife is albino and husband is normal. Let the dominant gene for normal skinned colour be denoted by A and that of albino by recessive a. The husband may be carrying the genes Aa with normal skin colour and also carrying an allele a for albino while the albino wife will have the genes aa. On coupling, the probability of the offspring Aa, 1 1 i.e., the gene A from husband and a from wife and that of aa is also normal skinned child is 2 2 i.e., the gene a from husband as well as from the wife. Suppose the first child is albino. The couple is interested to know the chance of the second child being albino. Since the skin colour of the first child has no bearing on the colour of the second child, the probability of the second child being albino 1 is also . 2 Now consider the problem in a different manner. The couple is interested to know the chance 1 and that of a normal skinned male birth. We know that the probability of the birth of a boy is 2 1 1 1 1 of a normal skinned child is . So the probability of a normal skinned boy is = and so 2 2 2 4 is the probability for a normal skinned girl. It mean that the couple may expect one normal skinned boy or girl out of four children.

1.4

ASCERTAINMENT OF HYBRID RATIOS

When a cross is made between a Yellow-Round (GGWW) and Green-Wrinkled (ggww) pea varieties, all the seeds in F1 are Yellow-Round (GgWw) as only GW type of gametes are produced by one parent and gw type by the other parent. F1 plants will produce four types of gametes (pollens as well as ova) and this will have sixteen types of zygotic combinations in F2 generation. The possible outcomes (zygotic combinations) are depicted in the chequer-board given below. Yellow-Round Green-Wrinkled (GG WW) (gg ww) Gametes GW gw Generation F1 will have gamete Gg Ww. However, in the F2 generation, individuals with combinations of pea colour and shape will appear as shown in the following chequer board. Parents

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STATISTICAL ANALYSIS OF QUANTITATIVE GENETICS

On counting, it is easy to note that the ratio of the four phenotypes namely, Yellow-Round, Yellow-Wrinkled, Green-Round and Green-Wrinkled is 9 : 3 : 3 : 1. The theoretical ratios of phenotypes can be obtained by considering the pairs of alleles separately and their ratio. Considering the pair of alleles individually, the ratio of Yellow to Green is 3 : 1 and for Round to Wrinkled is also 3 : 1. In the cross under consideration, it is apparent that when one character is fixed, the other character shows a clear cut ratio of 3 : 1. Therefore, it can be concluded that the characters under study are assorting (giving rise to different combinations) in an independent manner. In such a situation, the principle of inheritance is governed by Mendels second law known as the law of independent assortment. It states that Different characters in the offsprings of hybrids are distributed independently of each other. The expected ratio for a hybrid cross can be obtained by vectors multiplication as follows:

To show the ratio, elements of the two rows of the right hand matrix are written at a stretch and the ratio for GW, Gw, gW and gw turns out to be, 9 : 3 : 3 : 1, the same as obtained through the chequer board. In the same way, the expected ratio for a trihybrid cross obtained by considering the three pairs of genes through vector multiplication is as follows:

Writing the elements of the two rows of the right hand matrix at a stretch, we get the ratio as, 27 SGW : 9 SGw : 9 SgW : 3 Sgw : 9 sGW : 3 sGw : 3 sgW : 1 sgw

The above approach saves the labour of making the checker board.

Test of Goodness of F it Fit

A researcher is always anxious to confirm whether his frequencies in various classes follow the expected ratio or not. This test is known as a test of goodness of fit and is best performed by chi-square test. The test statistic is,
2

( Oi E i )2 Ei i =1
K

...(1.4.1)

Statistic 2 is distributed with (K 1) d.f. where K is the number of classes. Example 1.4.1. The data of Mendels experiment obtained in a dihybrid cross using green peas between Yellow-Round seeds and Green-Wrinkled seeds is reproduced here. Out of 556 seeds, the frequencies of seeds in four phenotypes were as follows:

BASICS OF GENETICS

11

Yellow-Round = 315, Green-Round = 108, Yellow-Wrinkled = 101, Green-Wrinkled = 32 We have to test the null hypothesis, H0 : The frequencies in four phenotypes follow the ratio of 9 : 3 : 3 : 1. H1 : H0 is not true. Expected frequencies are obtained as, E1 = E2 = E3 =
9 ! 313, 556 = 312. 75 = 16 3 ! 104 556 = 104. 25 = 16

3 ! 104 556 = 16 1 ! 35 556 = 34. 75 = E4 = 16

By the formula (10.4.1),

2 2 2 2 2 = ( 315 313) + (108 104 ) + (101 104 ) + ( 32 35) 313 104 104 35 = 0.0128 + 0.1538 + 0.0865 + 0.2571

= 0.5102 Tabulated value of for 3 d.f. and = 0.05 is 7.81. Since, the tabulated value of 2 is more than the calculated value, the null hypothesis is accepted. Thus, the experimental data supports the expected ratio.
2

Only one example is cited here just to impress upon the investigators that chi-square is a popular and powerful test in genetic theory to confirm the law of segregation. Probability is the base of most of the statistical methods. Sometimes one calculates probability directly or uses it indirectly. Now a number of statistical tools like path coefficient, coefficient of inbreeding, heritability, sire index, repeatability, general combining ability (g.c.a.), specific combining ability (s.c.a.), genetic and phenotypic correlations, analysis of variance, analysis of diallel crosses, stability models and many other methods meant for measuring genetic variation and relationships are dealt with adequately in the following discussion.

1.5

CROSSING OVER

As mentioned earlier, the genes are located on chromosomes in pairs and segregate independently at meosis. The two genes located closely on the same chromosome go together expressing simultaneously the characters of both genes in the offsprings. Such a situation is called linkage. The field trials conducted with characters of linked genes exhibited a few offsprings with recombination of characters. The phenomenon was explained by crossing over of a section of homologous chromosomes between two linked genes in a few cells. This was demonstrated histochemically in the dividing cells of gonads leading to formation of gametic cells. The chromosomes break at the point of crossing (chiasma) and join with the part of other chromatid thereby exchanging a section of homologous chromosomes and the genes located on it. The process results in recombination of genes.

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STATISTICAL ANALYSIS OF QUANTITATIVE GENETICS

Diagramatic representation of crossing over of homologous chromosomes.

Recombination Fraction
Consider the cross of two doubly homozygous populations AB/AB ab/ab under the assumption that they are linked. On crossing, F1 generation will be AB/ab. In case of complete linkage, the individuals AB/ab will produce two types of gametes AB and ab. Back cross i.e., crossing F1 with the recessive ab/ab, the progeny should have the type AB/ab or ab/ab only. But it has been observed that in some cases, the progeny have a few individuals of the type Ab/ab or aB/ab in addition to the type AB/ab or ab/ab. Though such an occurrence is seldom observed. Thus, the departure from usual independent segregation giving rise to the new genetic combinations is termed as recombination. At the same time, the genotypes are named as recombinants. The phenomenon of exchanging the genes between homologous chromosomes is known as crossing over. The proportion of recombinants occurring in the offspring due to crossing over is called recombination fraction and is denoted by 1 1 . The value of lies between 0 and , i.e., 0 . = 0 indicate complete linkage between 2 2 1 loci. But when = , one fails to detect whether there is no linkage or the loci are on different 2 chromosomes. It is apparent that each pair of homologous chromosomes belongs to a linkage group. Obviously, there are as many linkage groups as there are chromosomes in a genome. The phenomenon of linkage may be defined as, The tendency of two genes located on the same chromosome to be transmitted together in the process of inheritance. Consider a single locus with K alleles A1, A2, ... AK. In this situation there can be possibly K K homozygotes of the type Ai Ai (i = 1, 2, ...., K) and 2 heterozygotes of the type Ai Aj (i = 1, 2, ..., K and i < j). So the total number of genotypes is,
K +1 K(K 1) K(K + 1) ...(1.5.1) = = 2 2 2 For example, there are three alleles A, B and O in the blood group system ABO. A and B are

F I H K

K+

F KI H2 K

= K+

FG H

IJ K

dominant over O but are codominant to each other. So for three alleles, there will be

F 3 + 1I = 6 H 2K

genotypes. At the same time, the number of phenotypes can not be formulated as it depends on the

BASICS OF GENETICS

13

mode of dominance of alleles. In the present case, six genotypes and four phenotypes with three alleles A, B and O are, AA AO AB BB BO OO Genotypes Phenotypes A AB B O If we consider n loci where the ith locus contains Ki alleles, the total number of genotypes in respect of these n loci are,

F K + 1I F K + 1I ... F K + 1I = F K + 1I = 1 K ( K H 2 KH 2 K H 2 K H 2 K 2
n n 1 2 n i i =1 n i i =1

+1

...(1.5.2)

Now consider the case when organisms are polyploids say, 2m-ploids. Again suppose there are K alleles per locus and m homologous chromosomes, the number of genotypes is, = and the number of gametes is,

F K + 2 m 1I H K 1 K F K + m 1I H K 1 K F 5I = F 5I = 5 H 4K H 1 K
FG 3IJ = FG 3IJ = 3 H 2K H 1K
AAAa AAaA AaAA aAAA AAAa AAaa

F K + m 1I H m K F K + 2 m 1I H 2m K F 2 + 4 1I H 4 K
and the number of gametes is,

For instance, when m = 2, 2m = 4 i.e., tetraploid with two alleles A and a i.e., K = 2, the number of genotypes is, =

F 2 + 2 1I H 2 K
Combinations: Genotypes: AAAA AAAA

To elaborate further, for tetraploid all possible combinations of A and a are 24 i.e., 16. Genes A and a can occur in any manner on the four loci and the 16 combination result into 5 genotypes and 3 gametes as given below.

AAaa AaAa AaaA aAaA aaAA aAAa Aaaa aAaa aaAa aaaA Aaaa Whereas the three gametes are AA, Aa, aa. aaaa aaaa

1.6

STATISTICAL TESTS FOR SEGREGATION AND LINKAGE

Mendels first law of segregation is stated in section 1.2. On the other hand, linkage may be defined as, The tendency of two genes on the same chromosome to remain united in the process

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STATISTICAL ANALYSIS OF QUANTITATIVE GENETICS

of inheritance. The phenomenon of linkage is a sort of refutation of Mendels second law of independent assortment.

Statistical T est for Segregation Test

Consider heterozygous Aa type of individual. It can produce two gametes A and a. Different segregation ratios arise on the presumption that Aa produces the two gametes in equal number which can statistically be tested from the observed data. If we take its back cross with the recessive aa, it can result into two phenotypes Aa and aa in the ratio of 1:1. Suppose, an experiment or survey has a family of n individuals and it is observed that out of n, f are of the type AA and (n f ) are of the type aa. Now to confirm whether the segretation ratio of 1:1 holds good, make use of chisquare test provided n is sufficiently large. We know for two classes having the observed frequencies a and b for hypothetical ratio r : 1, the chi-square statistic is,
2 ( 1)

( a rb )2 r(a + b)
{ f 1.( n f )}2 1( f + n f )

...(1.6.1)

In the situation under consideration, a = f, b = (n f ) and r = 1. So the test statistic, = Suffix 1 of 2 indicates its d.f. If the calculated value of 2 is greater than the tabulated value of 2 for 1 d.f. and = 0.05 i.e., 3.841, One has to reject the segregation ratio 1:1. Rejection of segregation implies the presence of linkage. Consider now two loci with two factors A-a and B-b with dominance of A over a and of B over b. Let the observed frequencies in an experiment with double back cross Aa Ba ab ab be O1, O2, O3 and O4. The expected ratio of these frequencies in four phenotypic classes AB, Ab, aB, ab with no linkage between two factors will be 1 : 1 : 1 : 1. Let the total number of individuals under study is n. So be ...(1.6.2)

!o
i =1

= n. Under hypothetical ratio, the expected frequencies in each phenotype will

n . The validity of the expected ratio can easily be tested by 2-test. The test statistic in the 4

present case is,

2 = Statistic 2 has 3 d.f.

Fo nI H K ! 4
4 i i =1

n 4

...(1.6.3)

But if there is linkage between factors, the above situation does not prevail. We expect an excess of Aa Bb and ab ab if in F1 they were in coupling (AB/ab) and an excess of Ab ab or aB ab if they were in repulsion (Ab/aB). Thus the contrast Aa Bb + ab ab Abab aBab ...(1.6.4)

BASICS OF GENETICS

15

is suitable for detection of linkage. For single factor (A a) detection of linkage, the appropriate contrast is, Aa Bb + Ab ab aB ab abab Similarly, the contrast for single factor (B b) detection of linkage is, Aa Bb aBab Abab abab Above concept can be summarised in the table below. Table 1.6.1 Detection of linkage for backcrosses AaBb ab ab
Phenotypes AaBb AB Observed freq. Expected ratio Contrasts for linkage A vs. a B vs. b Linkage +1 +1 +1 +1 1 1 1 +1 1 1 1 +1 O1 1 Abab Ab O2 1 aBab aB O3 1 abab ab O4 1

...(1.6.5) ...(1.6.6)

It can easily be verified that the three contrasts given in the table are orthogonal. For testing of linkage in the three cases, there is the difference of the sum of frequencies of two phenotypes which are in the ratio 1:1. Therefore, the chi-square statistic each with 1 d.f. for testing of linkage and segregation are : For factor A a,
{( O1 + O2 ) ( O3 + O4 )}2 2 A a = n

...(1.6.7)

For factor B b, 2 B b = For linkage,

2 L

{( O1 + O3 ) ( O2 + O4 )}2 n {( O1 + O4 ) ( O2 + O3 )}2 n

...(1.6.8)

...(1.6.9)

Here, we test whether the families are homogeneous for two single factors A-a, B-b and for linkage. The decision about the hypothesis is taken in the usual way by comparing the calculated value of 2 with table value of 2 for 1 d.f. and level of significance. From chi-square table,
2 1 , .05 = 3.841.

The testing of segregation and linkage of two factors A a and B b for inbreeding experiments in doubly heterozygous individuals or in F2 generation can be done by chi-square test. For an intercross Aa Ba Aa Bb we have to test whether single factors A a or B b segregate in the ratio 3:1. Also the expected ratio of frequencies in the phenotypes A B , A bb,

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STATISTICAL ANALYSIS OF QUANTITATIVE GENETICS

aaB , aabb is 9 : 3 : 3 : 1. Without going into details, we give below the formulae for chi-square statistic each having 1 degree of freedom. For the factor A a, 2 A a = For the factor B b, 2 Bb = For linkage, 2 L =
{( O1 + 9 O4 ) 3( O2 + O3 )}2 9n {( O1 + O3 ) 3( O2 + O4 )}2 3n {( O1 + O2 ) 3( O3 + O4 )}2 3n

(1.6.10)

(1.6.11)

(1.6.12)

The decision about the expected ratio is taken in the usual manner. Example 1.6.1. In rabbits, two recessive genes produce a solid body colour and long hair respectively, in contrast to a spotted body colour and short hair which result from the dominant alleles. The results from a cross between the heterozygous spotted short haired rabbits and solid long haired rabbits are as follows : Class Spotted/short Spotted/long Solid/short Solid/long Frequency 48 5 7 40

If we want to perform a test for independent assortment in the above test cross to detect the linkage between the genes, we can proceed in the following manner. Denoting. Spotted/solid body = Ss Short hair/long hair = Ll Under independent assortment, the frequencies in the classes SL, Sl, sL, sl should be in the ratio, 1 : 1 : 1 : 1. Thus, H0 : Vs. H1 : 1:1:1:1 H0 is not true,

can be tested by chi-square test by the formula (1.6.3). Here n = 100. Hence,
2 2 2 2 2 = ( 48 25) + ( 5 25) + ( 7 25) + ( 40 25) 25 25 25 25

= 21.16 + 16.00 + 12.96 + 9.00 = 59.12. Tabulated value of for 3 d.f. and a = 0.05 is 7.81, which is less than the calculated value
2

BASICS OF GENETICS

17

of 2 = 59.12. So we reject H0. This leads us to conclude that there is no independent assortment and therefore, the genes S and L are linked. Example 1.6.2. In maize there is a dominant gene for coloured seed and another dominant gene for full seed. The recessive alleles of these genes produce colourless and shrunken seed. A homozygous coloured and full seeded maize plant was crossed with a colourless and shrunken seeded one i.e., F1 plants selfed to raise the F2 progeny resulted into the following phenotypic categories and their frequencies. Coloured and Full (CF) : Coloured and shrunken (Cf) : Colourless and full (cF) : Colourless and shrunken (cf) : O1 = 920 O2 = 310 O3 = 285 O4 = 85

On the basis of the above experimental data, one would like to verify whether: ( i ) The hypothetical ratio of 3:1 for phenotypic categories in F2 considering the segregation of only one character at a time stands true. (ii ) There exists a linkage between the genes for seed colour and shape. For the character C c, the hypothetical ratio of 3 : 1 can be tested by the chi-square test using the formula (1.6.10). Thus, 2 C c = Since
{920 + 310 3( 285 + 85)}2 3 1600

n = 1600 =
120 120 = 3.00 3 1600

2 Tabulated value of 2 for 1 d.f. and = 0.05 is 3.841, which is greater than C c = 3. 00. Hence we conclude that the genes pertaining to colour and colourlessness segregate in the ratio of 3 : 1.

Similarly for the characted F-f, the statistic chi-square by the formula (1.6.11) is, 2 Ff =

{920 + 285 3( 310 + 85)}2 3 1600 20 20 = 0.0833 3 1600

2 By comparing the value of 2 F f with 1( 0.05) = 3. 841, one can easily infer that the genes for full and shrunken traits segregate in the ratio of 3 : 1.

(iii) Now to perform a test for linkage between the genes for seed colour and shape, we make use of the chi-square statistic given by (1.6.12)
2 L =

{( 920 + 9 85) 3( 310 + 285)}2 9 1600

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STATISTICAL ANALYSIS OF QUANTITATIVE GENETICS

( 100 )2 = 0.6944 9 1600

2 Again the value of statistic is less than the tabulated value of 1(.05) . Hence, we accept the hypothetical ratio 9 : 3 : 3 : 1. This makes us to believe that there is no linkage between the genes responsible for seed colour and shape.

Overall chi-square for testing the agreement to the expected ratio 9 : 3 : 3 : 1 in the four categories can be worked out as follows : Expected frequencies in the four classes are 900, 300, 300 and 100 respectively. Thus, 2 =
( 920 900) 2 ( 310 300) 2 ( 285 300) 2 (85 100) 2 + + + 900 300 300 100

= 0.4444 + 0.3333 + 0.7500 + 2.2500 = 3.7777 = ! 3.78 2 Table value of for P = 0.05 and 3 d.f. is 7.815, which is greater than 3.78. Hence, it is concluded that the data supports the expected ratio. Notabene: It is interesting to verify that the sum of 2Cc, 2Ff and 2L is also 3.7777. It affirms that there is no linkage.

1.7 MULTIPLE-GENE HEREDITY OR ADDITIVE GENE EFFECTS

As early as 1909, Nilsson-Ehle conducting breeding experiments on wheat observed that crossing of Red and white varieties yielded hybirds of different intensities of red colour depending on the number of genes involved. He found that more the number of genes, darker the colour of seeds. He propounded that several genes were involved with only partial dominance and cumulative effect but individually indistinguishable so that each gene contributed to the redness of the seed. If the effects of two or more genes are a linear function of the individual gene effects, the effects of the genes are called multiple-gene heredity or additive gene effect. The condition is normally observed in quantitative traits like size of fruit, colour of seed, milk production etc. It is difficult to say whether each gene is contributing equally or differently. In the latter condition, the total effect Z will be a linear function of the gene effects. Consider K genes G1, G2, ..., GK located at different loci affecting the trait. Let the effect of the gene Gi be i (i = 1, 2, ..., K). then, Z = a11 + a22 + ... + aKK If i = for all i, then Z = (a1 + a2 + ... + aK) where i is the total number of genes. ...(1.7.2) ...(1.7.1)

1.8

INTERACTION OF GENES In the post Mendelian era, the breeding experiments revealed that the characters controlled by two or more genes located at different chromosomes do not express independently following the Mendelian laws of inheritence and dominance. Rather the presence of one type of genes interacts

BASICS OF GENETICS

19

with expression of other gene producing a new trait entirely different from both parents. Thus, a situation where one gene influences the expression of another gene controlling the same character is known as gene interaction. A classical example of gene interaction was demonstrated by Batson and Punnett (1906) in fowl. They selected fowls of two varieties, one with Rose comb and the other with Pea comb. The cross between these two varieties gave a new strain of comb called Walnut comb in F1. Interse cross between F1 yielded progeny in the proportion of 9/16 Walnut, 3/16 rose, 3/16 pea and 1/16 single type of comb. This is shown in the chequer board below : Rose comb RR pp F Gametes RP Rp rP rp RP [RRPP] [RRPp] [Rr PP] [Rr Pp] Rp [RRPp] (RRpp) [Rr Pp] (Rr pp) Pea comb PP rr

Rr Pp Rr Pp rP [RrPP] [RrPp] {rr PP} {rr Pp} rp [Rr Pp] [Rrpp] {rr Pp} rr pp Result [Walnut] 9/16 (Red) 3/16 {Pea} 3/16 single 1/16.

Earlier results had shown that either rose or pea when crossed with single comb fowl segregated independently in the ratio of 3:1 showing complete dominance over homozygous recessives. But the presence of even one dominant gene for red comb (R) and pea comb (P) together interacted with each other producing a new type called Walnut comb. However, double recessive (rrpp) expressed itself as single comb. In fact, the results do not defy the laws of inheritance but clearly exhibit the interaction between two dominant alleles and two recessive alleles.

Epistasis
This is another form of gene interaction. When two nonallelic genes affecting the same trait are present in an organism; the presence of one pair of genes masks or covers up the manifestation of other gene, then the phenomenon is called epistasis. The gene covering the expression of other gene is said to be epistatic while the gene masked is called hypotatic. In a particular strain of dog two different nonallelic genes say A and B were found to affect the colour of hairs. Gene A was responsible for black colour and presence of at least one B masked the colour resulting in white hair. Absence of A i.e., aa alongwith bb gave brown colour due to the absence of A and B. Thus, mating of AA BB (white) with aabb (brown) gave all white Aa Bb offsprings in F1 and a ratio of 12 : 3 : 1 for white, black and brown in F2 respectively as depicted in the crosses below.

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STATISTICAL ANALYSIS OF QUANTITATIVE GENETICS

Gametes AB Ab aB ab F2

AB AABB White AABb White AaBB White Aa Bb White White 12

Ab AABb White AAbb Black AaBb White Aa bb Black Black 3

aB AaBB White AaBb White aaBB White aaBb White Brown 1

ab AaBb White Aabb Black aaBb White aabb Brown

The presence of one B has masked the colour of hair. So B is epistatic to A and A is hypostatic. This is an example of dominant espistasis. Similarly there are several types of epistatic effects such as recessive epistasis giving a ratio of 9 : 3 : 4, incomplete duplicate recessive yielding a ratio of 9 : 6 : 1, duplicate recessive epistasis resulting in the ratio of 9 : 7, duplicate dominant epistasis yielding the ratio of 15 : 1 and dominant and recessive epistasis showing the ratio of 13 : 3 in F2. All these can be verified by the readers if needed at all.

QUESTIONS AND EXERCISES

1. What is the constitution of a cell and its function? 2. Differentiate between somatic and gametic cells. 3. Write the conditions that are known as diploid, triploid, haploid and polyploids. 4. Classify homogametic and heterogametic conditions. 5. What do you understand by a gene and its alleles? 6. What kind of activity takes place under mitosis and meiosis? 7. Name the scientist who is called as father of genetics and why? 8. Enunciate Mendles laws of genetics. 9. Distinguish between genotypic and phenotypic variations. 10. How does laws of probability help in heredity determinations? 11. What is hybridization and how one can ascertain the hybrid ratios theoretically? 12. Explain chi-square test for multiple ratios. 13. What is meant by crossing over in genetic theory? 14. Define and explain the term recombination fraction. 15. Explicate the terms epistatic and hypostatic. 16. An experiment on chlorophyll inheritance in maize consisted of 1092 heterozygous self fertilized green plants which segregated into 869 dominated green plants (DG) and 223 recessive yellow plants (dg). The theoretical ratio of DG to dg is 3 : 1. Test statistically whether the experimental data conform the theoretical ratio?

BASICS OF GENETICS

21

17. Following phenotypic frequencies were obtained in an experiment from a F2 population. Phenotypes : Frequency : AB 475 Ab 122 aB 128 ab 75

The expected ratio of frequencies in the given four classes is 9 : 3 : 3 : 1. (i) Test whether the observed frequencies are in agreement with the expected ratio at 1 per cent probability. (ii) Test in case of F2 under back cross between two factors (A, a) and (B, b) for linkage and segregation. (iii) Test for segregation and linkage between two factors (A, a) and (B, b) in case of back cross in F1. 18. The following data were observed for hybrids of Datura. Flowers violet, fruits prickly Flowers violet, fruits smooth Flowers white, fruits prickly Flowers white, fruits smooth 47 12 21 3

Using chi-square test, find the association between colour of flowers and character of fruit at 5 per cent level of significance. 19. What are autosomes? Write the number of autosomes in human beings. 20. What do you understand by linkage and crossing over? 21. Define genic interaction and give its example. 22. How can we say that additive gene effect exists in a progeny? 23. What is the chemical composition of a chromosome and a gene? 24. Name one most important difference between somatic and gametic cells.