EXPERIMENT: 1

PLANT BREEDER’S KIT

Equipments Uses
1) Scissor
It is required for removal of unwanted
leaves, buds, etc.

2) Needle

Long and pointed needle is required to

open the small flower buds.

3) Forceps/Tweezers

Fine forceps with long point is required

for removing the anthers during
emasculation and to transfer the pollen
grains from male flowers to the stigma of
emasculated female flowers

4) Brush
Hair brush is required for collecting the
pollen grains and dusting on the stigma of
emasculated female flowers.

5) Spirit bottle
A small bottle of alcohol or spritis needed
to sterilize the scissors, needles, forceps
and brush as well as hands during the
crossing work

6) Magnifying Lens It is useful in emasculating small flower

buds and it is also required to observe the
stigma of emasculated flower to confirm

1
 that stigma does not carry any anthers or
pollen grains

Bags Bags are required to cover the flowers or

7)
inflorescences prior to pollination and
after the pollination. Bags are of different
kinds and sizes are used depending upon
types of crops and size of flowers, viz.,
brown paper bag, muslin cloth bag,
parchment paper bag, etc.

8) Labels

Paper or aluminum labels are used to

write the necessary information. The
information on the label should be written
with non-copying pencil by hands

9) U - Pins or Wire rings

They are used for holding the paper bag

kept on the flowers or inflorescences.

10) Field Notebook All the important observations required to

be takenduring the breeding works are
written in the field notebook. This note-
book should have the information/record
of number of crosses, date of selfing or
emasculation, flowering, maturity,
number of seeds set in crossed flower and
about the plant selected in segregating
generations
Assignment: List the various apparatus of breeder's kit and write its uses.

2
 EXPERIMENT: 2

GERMPLASM AND ITS CONSERVATION

PLANT GENETIC RESOURCES

The sum totals of hereditary material i.e. all the allele of various genes, present in a crop
species and its wild relatives is referred to as germplasm. This is also known as plant
genetic resources or genetic stock. Germplasm consists of the following five types of
materials:

1. Land Races
These are locally adapted, traditional varieties, which had evolved over centuries through
both natural and artificial selection, but without systematic and sustained plant breeding
efforts. Since, they are sources of many valuable genes, their conservation is essential.
The main drawbacks of land races are that they are less uniform and low yielders. Land
races were first collected and studied by N.I Vavilov in rice.

2. Obsolete Cultivars
Improved varieties of recent past are known as obsolete cultivars. These varieties were
developed by systematic plant breeding efforts and popular earlier but now have been
replaced by new varieties.

3. Modern Cultivars

The currently cultivated varieties are referred to as modern cultivars. These varieties have
high yield potential and uniform as compared to obsolete varieties and land races.

4. Wild Form

Wild forms are the wild species from which crop species were directly evolved.

3
GERMPLASM CONSERVATION

Germplasm conservation: - It is defined as the maintenance of germplasm of a

species by in-situ conservation, field collection, seed banks, slow growth cultures,
cryopreservation and DNA banks.

Genetic erosion: - The gradual loss of variability in the cultivated forms and in
their wild relatives is referred to as genetic erosion.

The very objective of germplasm conservation is to preserve the genetic diversity of a

particular plant or genetic stock for its use at a time in future.

Organizations associated with germplasm

A. International level

IPGRI – International Plant Genetic Resources Institute, Rome (Italy)

B. National level
1. NBPGR – National Bureau of Plant Genetic Resources: - It deals with
quarantine inspection of Agri. and Horti. Crops, New Delhi
2. FRI – Forest Research Institute: - It deals with quarantine inspection of forest
trees, Dehradun
3. BSI – Botanical Survey of India: - It deals with quarantine inspection of
medicinal and other plant sps. Kolkata

Methods of germplasm conservation

1. In situ conservation 2. Ex situ conservation

In situ conservation
1. On site conservation.
2. Conservation of germplasm under natural habitat is referred to as in situ
conservation.

4
 3. This is achieved by protecting this area from human interference. Ex. Natural
park, Biosphere reserve and Gene sanctuary etc.
4. A gene sanctuary not only conserves the existing genetic diversity present in the
population, it also allows evolution to continue. As a result, new alleles and new
gene combinations would appear with time.
5. In situ conservation has practical limitations associated with shrinking of natural
habitats, urbanization, industrialization and changing government policies

Merits:

 In this method of conservation, the wild species and the complete natural or semi-
natural ecosystems are preserved together.
Demerits:

 Each protected area will cover only very small portion of total diversity of a crop
species, hence several areas will have to be conserved for a single species.
 The management of such areas also poses several problems.
 This is a costly method of germplasm conservation.

Ex situ conservation

1. Off site conservation.

2. Conservation of germplasm away from the area of its natural habitat. Ex. Seed
gene banks, Plants or Field bank, Cell or Organ bank, DNA gene bank, herbal
gardens, in vitro repositories etc.
3. Cheap method of germplasm conservation.
4. Handling of germplasm is easy. It is possible to preserve entire genetic diversity of
a crop species at one place.
5. Preservation in the form of seed is the most common and easy method, relatively
safe, requires minimum space and easy to maintain.

5
 This type of conservation can be achieved in the following five ways.

1) Seed banks:
Germplam is stored as seeds of various genotypes. Seed conservation is quite easy,
relatively safe and needs minimum space. Seeds are classified, on the basis of their
storability into two major groups.

1) Orthodox and 2) Recalcitrant

Orthodox seeds: Seeds of this type can be dried to low moisture content of 5% and
stored at a low temperature without losing their viability are known as orthodox seeds.
(eg.) Seeds of corn, wheat, rice, carrot, papaya, pepper, chickpea, cotton, sunflower.

Recalcitrant: Seeds which show very drastic loss in viability with a decrease in moisture
content below 12 to 13% are known as recalcitrant seeds. (e.g) citrus, cocoa, coffee,
rubber, oil palm, mango, jack fruit etc.

Seed storage: Based on duration of storage, seed bank collects are classified into three
groups. (1) Base collections. (2) Active collections and (3) Working collection.

Base collections: Seeds can be conserved under long term (50 to 100 years), at about -
20OC with 5% moisture content. They are disturbed only for regeneration.

Active collection: Seeds are stored at 0OC temperature and the seed moisture is between
5 and 8%. The storage is for medium duration, i.e., 10-15 years. These collections are
used for evaluation, multiplication, and distribution of the accessions.

Working collections: Seeds are stored for 3-5 years at 5-10OC and the usually contain
about 10% moisture. Such materials are regularly used in crop improvement
programmes.

2. Plant Bank: (Field or plant bank) is an orchard or a field in which accessions of fruit
trees or vegetatively propagated crops are grown and maintained.

6
Limitations:

1. Require large areas

2. Expensive to establish and maintain
3. Prone to damage from disease and insect attacks
4. Man – made
5. Natural disasters
6. Human errors in handling

3. Shoot tip banks: Germplasm is conserved as slow growth cultures of shoot-tips and
node segments. Conservation of genetic stocks by meristem cultures.

4.Cell and organ banks: A germplasm collection based on cryopreserved (at – 196OC
in liquid nitrogen) embryogenic cell cultures, somatic/ zygotic embryos they be called
cell and organ bank.

5.DNA banks: In these banks, DNA segments from the genomes of germplasm
accessions are maintained and conserved.

Activities in germplasm collections and conservations

I. Collection
II. Introduction
III. Exchange
IV. Evaluation
V. Documentation
VI. Safe conservation
VII. Sustainable management of germplasm

7
Germplasm Documentation

Each germplasm accession is given an accession number. This number is pre fixed in
India, with IC (Indigenous collection), EC (exotic collection) or IW (Indigenous wild).

CRYOPRESERVATION

Cryopreservation is the storage of viable biological material at ultra-low temperatures,

which provides a means for the long-term stable storage of plant germplasm.
Cryopreservation at -196ºC in liquid nitrogen (LN) has been considered to be an ideal
tool which offers long-term storage capability, maximal stability of phenotypic and
genotypic behavior of stored germplasm, and minimal storage space and maintenance
requirements.

THE GENE POOL SYSTEM OF CLASSIFICATION

Gene pool of a crop included all cultivars (Obsolete and current), wild species and wild
relatives containing all the genes available for breeding use.

Based on degree of relationship, the gene pool of a crop can be divided into three
groups.

These are briefly discussed below:

1. Primary Gene Pool (GP1):

The gene pool in which intermating is easy and leads to production of fertile hybrids is
known as primary gene pool. It includes plants of the same species or of closely related
species. In such gene pool, genes can be exchanged between lines simply by making
normal crosses. This is also known as gene pool one (GP1). This is the material of prime
breeding importance.

8
2. Secondary Gene Pool (GP2):

The genetic material that leads to partial fertility on crossing with GP1 is referred to as
secondary gene pool. It includes plants that belong to related species. Such material can
be crossed with primary gene pool, but usually the hybrids are sterile and some of the
progeny to some extent are fertile. Transfer of gene from such material to primary gene
pool is possible but different. This type of gene pool is also known as gene pool two
(GP2).

3. Tertiary Gene Pool (GP3):

The genetic material which leads to production of sterile hybrids on crossing will primary
gene pool is termed as tertiary gene pool or gene pool three (GP3). It includes material
which can be crossed with GP1, but the hybrids are sterile. Transfer of gene from such
material to primary gene pool is possible with the help of special techniques.

Assignment:
1. Write in brief different methods of germplasm conservation.
2. Explain gene pool concept and its group.

9
 EXPERIMENT: 3

MODE OF POLLINATION

POLLINATION AND ITS TYPES

Pollination: Pollination is defined as the transfer of pollen grains from anther to the
stigma is called as pollination.

Types of pollination: -
1. Self pollination

2. Cross pollination

1) Self pollination: -

It is refers to the pollen from an anther may fall on to the stigma of the same flower is
called as self-pollination or autogmay.

2) Cross pollination: -

It refers to the pollen grains from flower of one plant are transmitted to the stigma of
flowers of another plant it is called as cross-pollination or allogamy.

3) Geitonogamy: -

When pollen from a flower of one plant falls on the stigmas of other flowers of the
same plant. e.g. maize.

1) SELF POLLINATION: -

o Development of seed by self-pollination is known as autogamy or self-pollination.

o Auotogamy is the closest form of breeding.
o Auotogamy leads to homozygosity.
o Such species develop homozygous balance & do not exhibit significant inbreeding
depression.

10
 o But in most of these species, self pollination is not compete & cross-pollination
may occur up to 5%.
o The degree of cross-pollination in self-pollinated species is affected by several
factors. e.g. variety, environmental condition like temperature, humidity &
location.

MECHANISM PROMOTING SELF POLLINATION

There are various mechanisms that promote self-pollination, which are generally
more efficient than that cross-pollination.

Bisexuality: - Presence of male & female organs in the same flower is known as
bisexuality. The presence of bisexual flowers is a must for self-pollination. All the self-
pollinated plants have hermaphrodite flowers. The various mechanisms which promote
self pollinations are as under

1) Homogamy: -
 Maturation of anthers and stigma of a flower at the same time is called as
homogamy.
 As rule, homogamy is essential for self-pollination.

2) Cleistogamy: -
 When pollination and fertilization occur in unopened flower bud is called as
cleistogamy.
 It ensures self-pollination prevents cross-pollination.
 In th is case flowers do not open at all this ensures complete self pollination since
foreign pollen cannot reach the stigma of a closed flower e.g. wheat, oats, barley
and other grasses.

4) Chasmogamy: -

 In plants, fertilization after opening of flower is known as chasmogamy.

11
  It promotes self-pollination. e.g. wheat, barley, rice, oats etc.

5) Position of anthers: -
 In some species, stigma are surrounded by anthers in such a way that self-
pollination is ensured.
 Such situation is found in tomato and brinjal.
 Some legumes, the stamens and stigma are enclosed by the petals.
e.g. green gram, black gram, soybean, chickpea and pea etc.

GENETIC CONSEQUENCES OF SELF POLLINATION

 Self-pollination leads to a very rapid increase in homozygosity.

 Therefore populations of self-pollinated species are highly homozygous.
 Self-pollinated species do not show inbreeding depression, but may exhibit
considerable heterosis.
 Therefore, the aim of breeding methods & develop homozygous varieties.

2) CROSS POLLINATION:-

 Development of seed by cross-pollination is called as allogamy.

 In cross pollination species the transfer of pollen from a flower to the stigmas of
the others.

May brought about by

1) Wind (anemophily)
2) Water (hydrophily)
3) Insects (anemophly)

Many of the crop plants are naturally cross-pollinated. In many species a small amount
(up to 5%) of selfing may occur.

MECHANISM PROMOTING CROSS POLLINATION:

There are several mechanisms that facilitate cross-pollination

12
I) Dicliny: -
It refers to unisexual flowers. The flowers are either staminate (male) or pistilate
(female).

This is of two types: viz. 1) Monoecy and 2) Dioecy

1) Monoecy: -
 Staminate & pistilate flowers occur in the same plant, either in the same
inflorescences. e.g. castor, mango, banana, coconut, or in separate
inflorescences, e.g. maize.
Other monoecious species are cucurbits, walnut, chestnut, strawberries, rubber,
grapes & cassava.

2) Dioecy: -
 The male & female flowers are present on different plants, i.e. the plants in such
species are either male or female, e.g. papaya, datepalm, hemp, asparagus, &
spinach.
 In general the sex governed by a single gene, e.g. asparagus & papaya. There are
hermaphrodite plants in condition to male & female plants, & a number of
intermediate forms may also occur.

II) Dichogamy: -
Stamens and pistils of hermaphrodite flowers may mature at different times. There are
of two types

a) Protogyny – pistils mature before stamens. e.g. bajara

b) Protandry - stamens mature before pistils.e.g, maize and sugarbeet.

III) Heterostyly: -
When styles and filaments in a flower are different length, it is called heterostyly.
e.g. linseed.

13
IV) Herkogamy: -
Hindrance to self-pollination due to some physical barriers such as presence of
hyline membrane around the anther is called as herkogamy.eg alfalfa.

V) Self incompatibility: -
It refers to the failure of pollen from a flower to fertilize the same flower or other
flowers on the same plant.

There are two types of self-incompatibility:

a) Sporophytic incompatibility: -
Self-incompatibility is controlled by the genotype of the pollen producing plant is
called as sporophytic incompatibility, e.g. cabbage, cauliflower, radish.

b) Gametophytic incompatibility: -
Self-incompatibility is controlled by the genetic constitution of pollen is called as
gametophytic incompatibility. e.g. potato tomato, rice.

VI) Male sterility: -

 It is refers to the absence of functional pollen grains in otherwise hermaphrodite
flowers is called male sterility.
 Male sterility is not common in natural populations. But it is great value in
hybrid seed production.
 It prevents self-pollination and promotes cross-pollination.

 There are following three types of male sterility:

a) Cytoplasmic male sterility: -

The pollen sterility which is controlled by cytoplsamic genes or plasma genes is
called as cytoplsamic male sterility. e.g. onion .

14
 b) Genetic male sterility: -
The pollen sterility, which is controlled by nuclear genes, is termed as genic or
genetic male sterility. e.g. barley , maize, wheat , cotton , sorghum Lucerne,
cucurbits, tomato and sugarbeet.

c) Cytoplasmic genetic male sterility: -

When pollen sterility, which is controlled by both cytoplamic and nuclear genes,
is known as cytoplasmic genetic male sterility. E,g wheat maize , sorghum , rice,
sunflower cotton, tobacco, tomato, onion, sugarbeet , carrot linseed.

GENETIC COSEQUENCES OF CROSS POLLINATION

 Cross pollination preserves and promotes heterozygosity in a population

 Cross-pollinated species are highly heterozygous.
 The breeding method in such a species aim at improving the crop species without
reducing heterozygosity to an appreciable degree.
 Usually, hybrid or synthetic varieties are the aim of breeder wherever the seed
production of such varieties is economical feasible.

3) OFTEN CROSS-POLLINATED SPECIES

 Cross-pollination often exceeds 5% and may reach upto 30% such a species is
known as often cross-pollination. e.g. jawar cotton, arhar , safflower , etc.

DETERMINATION OF THE MODE OF POLLINATION

1) The first step – Determine the mode of pollination of species is to examine its
flower. Mechanism like dioecy, protogyny, protandry, cleistogamy are easily
detected; they clearly indicate the mode of pollination.

2) The second step: -

 Isolate single plants and recording seed set under isolation.
 Space isolation, i.e individual plants grown at sufficient distance to prevent
cross-pollination, is preferable to isolation by bags or cages.

15
  Isolation of bags and or cages may create an environment unfavorable for
pollination and seed set.
 Failure of set seed in isolation proves the spaces to be cross-pollinated.
 However, setting of seed s only indicates of self-pollination.

Finally, the effect of selfing (inbreeding) on the vigour of plants should be studied.
Loss in vigor due to inbreeding is common in cross-pollination, but self-pollinators
show to inbreeding depression.

DETERMINATION OF THE AMOUNT OF CROSS POLLINATION

The amount of cross-pollination is determined by planting two strains of the

concerned species in a mixed strain.

1) One strain is homozygous for a dominant character preferably an easily

recognizable seed or seedling character.
2) Other strain has the recessive form of the character.
3) The two strains are planted in such a manner that each plant of recessive strain is
surrounded by plants of the dominant strain to provide abundant pollen.
4) The seeds from the plants of only recessive strain are harvested.
5) The percentage of seeds carrying the dominant allele represents the percentage of
crosspollination in the species.
6) The frequency of cross-pollination varies greatly with variety, weather condition.

e.g. In a study on safflower, the estimates of out crossing in different varieties grown
in the same time at the same location from 0-8.7% similarly, the amount of cross
pollination in a single variety grown at a several locations varied from 1.3-9.8%.
Therefore such a study should include several varieties of the crop and the study
should be conducted at several locations for two more years.

16
DETERMINATION OF THE AMOUNT OF CROSS – POLLINATION
Crop : Castor Inbred / Varieties:1.VI – 9 Green Stem (Recessive)

2. 48–1 Red Stem (Dominant)

D D D
D
D D D
D D D
r D
D D r
r D
D D D
D
D D
D D D D
D

Steps: 1. Sowing: Where, r = VI-9 and D = 48-1

0 0 0

0 0 0 00 0

000 0 0 0 0 0 00 0 0

No. of Plants 100 No. of Plants 100 No. of Plants 100

Plot No.1 Plot No.2 Plot No.3

Recording of Observations:

Character Plot Plot Plot Average

No.1 No.2 No.3

Green Stem Plant 55 60 53 -

(Recessive
Red Stem Plant 45 40 47 -
(Dominant)
% Cross 45 40 47 44
Pollination
The average of cross-pollination is 44 %.

17
SIGNIFICANCE OF POLLINATION

1. Gene action.
2. Genetic constitution
3. Adaptability
4. Genetic purity
5. Transfer of genes

1. GENE ACTION: -
In refers to mode of expression of genes for various characters in a population. Gene
action is of two types:

1) Additive gene action: - It is associated with homozygosity and is more in self-

pollination species.
2) Non additive gene action or (dominant and epitasis): - It is associated with
heterozygosity and therefore is more cross-pollinated species.

2. GENETIC CONSTITUTION: -
The breeding population is of four types viz. heterozygous, homozygous,
homogeneous, and heterogeneous.

1) Heterozygous: - Individuals have dissimilar alleles at the corresponding loci and

are pure types.
2) Homozygous: - Individuals have dissimilar alleles at the corresponding loci and
are of pure types. (They do not segregate on selfing)
3) Homogeneous: - The genetically similar population is known as homogeneous.
4) Heterogeneous: -
 Genotypically dissimilar population it is known as heterogeneous.
 It may again the homozygous or heterozygosity and cross-pollination results in
heterozygosity because in breeders have advantageous of homozygosity and
outbreeders have advantage of hererozygosity.

18
3) ADAPTABILITY:
It refers to stable performance of a variety over a wide range of environmental
conditions.

 The cross pollinating species have better adaptability (Buffering capacity) than
self pollinating species, because of more heterozygosity and heterogeneity in
the former case.
 The heterozygosity populations have broad genetic base. Such a populations
have greater capacity to stabilize productivity over a wide range of changing
environments.
 Heterogeneous population gives more stable yields over similarly;
heterozygous individuals such as F1 hybrids are more stable to environmental
variations than their homozygous parents.
 Thus mode of pollination plays a key role in varieties adaptability.

GENETIC PURITY: -

 Self-pollination maintains the genetic purity of a species and the purity of a

variety of a self-pollinating crop can be easily maintained for several years.
 Cross-pollination creates heterozygosity and genetic composition of a cross-
pollinating variety change gradually.
 Hence, self-pollination is essential even in cross-pollinated species to maintain
the purity in parental lines.

5) TRANSFER OF GENES:

Cross-pollination permits transfer of desirable genes from one species to another. Or

combining of desirable genes from different sources in to a single genotype is
possible through cross-pollination.

19
Classification of crop plants based on mode of pollination and mode of reproduction

Mode of pollination and Example of crops

reproduction
A. Autogamous species Wheat, rice, barley, oats, chickpea, pea,
1. Seed propagated cowpea, lentil, green gram, black gram,
soybean, common bean, moth bean,
linseed, sesamum, khesari, sunhemp,
chillies, brinjal, tomato, okra, peanut.

B. Allogamous species Corn pearlmillet, rye, Alfalfa, radish,

1. Seed propagated cabbage, sunflower, sugarbeet, castor,
red clover, white clover, safflower,
spinach, onion, garlic, turnip, squash,
muskmelon, watermelon, cucumber,
pumpkin, oilpalm, carrot, coconut,
papaya.

Sugarcane, coffee, cocoa, tea apple,

2. Vegetatively propagated peaches, pears, cherries, grapes,
almond, strawberries, pine apple,
banana, cashew nut, rubber, sweet
potato, etc.
C. Often-allogamous species. Sorghum, cotton, triticale, pigeonpea,
(Seed propagated) Tobacco.

Assignment:

1. Explain in brief different modes of mechanism which promote self and cross
pollination in crop plants
2. How the mode of pollination in crop plant can be determined?
3. Classify different agricultural crops on the basis of their mode of pollination.

20
 EXPERIMENT: 4

ESTIMATION OF HETEROSIS AND INBREEDING DEPRESSION

The heterosis term was given by Shull in 1914. The heterosis may be defined as the
superiority or inferiority of an F1 hybrid over both its parents in terms of yield and some
other characteristics.

Estimation of different types of heterosis – 3 types

1. Superiority of F1 over mid parental value is known as average Heterosis

– Heterosis (%) =[ (F1 – MP)/MP] x 100

o F1 = F1 mean
o MP = mid parental value

2. Superiority of F1 over better parental value is known as heterobeltiosis

– Heterobeltiosis (%) =[ (F1 – BP)/BP] x 100
o F1 = F1 mean
o MP = mid parental value

3. Superiority of F1 over standard check or commercial variety is known as

economic or standard heterosis. This type of heterosis has commercial importance
– Standard Heterosis (%) =[ (F1 – SC)/SC] x 100
o F1 = F1 mean
o SC : mean value of standard check

INBREEDING

 Mating between individuals related by descent or ancestry is known as inbreeding

OR
 Mating between closely related individuals like brother sister mating or sib
mating

21
Inbreeding depression – The severe reduction or loss in vigour and fertility due to
inbreeding is known as inbreeding depression

OR

The reduction or loss in vigor and fertility as a result of inbreeding.

Estimation of inbreeding depression

Since the maximum decline is reflected in F2 generation, the inbreeding depression can
be computed by relative data on F1 and F2 for any character as under

Inbreeding Depression (%) = [ F2-F1/ F2 ] x 100

Characteristics

 Concept of inbreeding depression was given by East and Shull (1909)

 Marriages between individuals related by ancestory are restricted in early times in
Hindu society
 Inbreeding depression is more common in cross pollinated crop
 The chief effect of inbreeding is increase in homozygosity in the progeny

Effects of inbreeding

 Appearance of lethal and sub lethal alleles

o Lethal – leading to death
o Sub lethal and sub vital – reduce survival and reproduction rate
o Example – rootless seedling, defects in flower structure, chlorophyll
deficiency
 Reduction in vigour
 Reduction in reproductive ability
 Separation of the population into distinct lines
 Increase in homozygosity
 Reduction in yield

22
Degree of inbreeding depression

1. High inbreeding depression

– Alfalfa, carrot
– Selfing produced lethal characteristics
– > 25 % yield reduction
2. Moderate inbreeding depression
– Maize, Jowar, Bajra
– Proportion of lethal characteristics are relatively very low
3. Low inbreeding depression
– Onion, cucurbits, sunflower, rye
– Smaller proportion of lethal characteristics
4. No inbreeding depression
– Self-pollinated crops, showing homozygous balance

The differences between inbreeding and heterosis

Inbreeding depression Heterosis
(1) Results from mating between (1) Results from crossing
closely related individuals between unrelated strains
(2) Inbreeding depression is the (2) Heterosis is the increase in
decline in fitness and vigour fitness and vigour with
with decreased heterozygosity increased heterozygosity.
(3) Inbreeding depression results (3) Heterosis the unfavourable
due to fixation of unfavourable recessive genes of one line
recessive genes in F2. (parent) are covered by
favourable dominant genes
of other parent.

23
Assignment:

1. What is heterosis, heterobeltiosis and economic heterosis? How will

you calculate these?
2. Write the features of heterosis.
3. What are the factors affecting heterosis?
4. What is inbreeding ? Write effects of inbreeding.
5. What is inbreeding depression? Describe in brief the degrees of inbreeding
depression

24
 EXPERIMENT: 5

EMASCULATION AND HYBRIDIZATION

WHAT IS HYBRIDIZATION?

The mating or crossing between two plants or lines having dissimilar (unlike)
genetic constitution (genotypes) is called hybridization. OR Hybridization is an
artificial controlled pollination in which pollen grains of desired male parent is
transferred to the stigma of the desired female parent.

 Objectives of hybridization:

1. To create genetic variability.

2. To combine the desirable character (s) into a single plant.
3. To know the general combining ability of the parents and the
specific combining ability of the crosses.
4. To study the pattern of inheritance of the characters.
5. To exploit and utilize the hybrid vigor.
6. To produce hybrid varieties.

 TYPES OF HYBRIDIZATION:

 In hybridization, plants or parents which are to be crossed may belong to

the same species, different species or different genera.

There are four types of hybridization which are as under.

1. Inter varietal/Intra specific hybridization:

 The parents involved in hybridization belong to be same species.

 They may be two strains or varieties or races of the same species.
 This is the most common type of hybridization used in crop improvement
and have produced many varieties in different crop plants.

25
 e.g. In bread wheat – a cross between GW 496 and GW 503.
In groundnut – a cross between GG-2 and GAUG 10; and
GG 11 and GAUG10

2. Inter specific / Intra generic hybridization:

 Crosses are made between the plants of two different species of the same genus.
In other word, the crossing between two species of the same genus is called inter
specific hybridization.
e.g. Triticum aestivum x T. durum;
Oriza sativa x O. perennis;
Gossypium hirsutum x G. arborium;
Nicotiana tabacum x N. rustica, etc.
 This type of hybridization is used to transfer one or few simply inherited
characters like disease/pest resistance or drought resistance from wild species
into cultivated species.

3. Inter generic hybridization:

 A cross is made between the plants belonging to two different genera. It is used
to transfer desirable characteristic from one genus to another.

e.g. Triticum spp x Secale cereal

Triticale

(First man made artificial cereal)

 Inter-specific and inter-generic hybridization are also known as wide crosses or

distant crosses (distant hybridization) as parents are distinctly related.

26
 4. Introgressive hybridization:

 It is one kind of inter-specific hybridization. In this type of hybridization hybrids

may have repeatedly backcrossed to one of the parental species.
 Thus most of genotype of that parental species would be recovered along with
few genes from the other parental species.

e.g. Primitive maize x Tripsacum

Modern maize

 Precautions to be taken during hybridization:

1. The plants in which the emasculation is to be done should be enough

apart from other plants.
2. A proper bud size should be selected for the emasculation.
3. Sterilized instruments should be used for emasculation.
4. Do not injure the floral parts.
5. Anthers should be removed by holding their filaments with forceps.
6. The emasculation should always be done at the time when stigma is not
receptive.
7. Magnifying lens may be used to confirm whether there is presence of
any piece of anthers on emasculated flower bud.
8. Sufficient number of buds should be emasculated on a branch to
minimize the chance of failure of across.
9. Un-emasculated buds, opened flowers, disease/insect damaged flowers
and fruits on the branches should be removed before bagging.
10. The label and tags should be light in weight so that flower or

27
 inflorescence may not be bending.

 Important features of hybridization technique:

1. Make careful study of the structure of flower before commencing

operations. This may be with or without the aid of a dissecting microscope.
2. Determine which flowers produce larger, better seeds and which set seeds
most freely.
3. Learn the normal time and method of blooming of the flowers and the
length of time that the pistil will remain receptive and the pollen grains
capable of functioning.
4. Procure the necessary instruments and see that these are of an efficient
kind for the work to be undertaken.
5. Be careful not to injure the following parts any more than necessary. Do
not remove the surrounding flower parts i.e. petals, glumes, flag leaf, etc.
unless required.
6. A few crosses carefully made are of much greater value than many done
with less care.

Steps/procedure of hybridization:

1) Selection of parents

(i) The parents must be homozygous.

(ii) Desirable character(s) must be present in the parents.

(iii) If possible, female parent should be prevailing or ruling superior

agronomical variety.

2) Selection of flowers

(i) Select immature and healthy flowers prior to anthesis.

28
 (ii) Remove all other flowers keeping only few flowers for hybridization.

3) Preparation of female flower

(i) The preparation of female flower is called emasculation, in which all the anthers
are removed from the female flower prior to anthesis.
(ii) Remove all other unwanted leaves, buds, and other vegetative parts.
(iii) In case of dicliny or dioecy plant/flower, emasculation is not necessary but the
flower buds are to be protected from foreign pollens prior to anthesis.
(iv) Various methods of emasculation are being utilized in hybridization programme
which are asunder.

 METHODS OF EMASCULATION:

(A) Hand emasculation:

(i) Remove all the anthers by forceps or pointer. Take care that even a piece of
anther should not remain in the emasculated female flowers.
(ii) Usually emasculation is to be started from top flower and proceed downward.
This practice will avoid the falling of anther on emasculated female flowers.
(iii) Emasculation is generally done in late evening or early morning, one day before
the anthers are expected to mature and stigma become fully receptive.

(B) Temperature treatment:

The removal of anthers by forceps or pointer is very laborious work for the crop
plants with very small flowers. Hence, temperature (hot or cold water) isused.

(i) Hot water treatment:

 Hot water has been used to kill pollen in sorghum, rice, grasses, etc. The
flowers are immerged in hot water with temperature ranging from 45ºC to 53ºC
for 5-10 minutes, depending upon the species.

29
 (ii) Cold water treatment:

 Chilling treatment (cold water) has been used in wheat and rice with
temperature around freezing.
 Use of hot or cold water is a simple procedure since a thermo-flask is used to
maintain desire temperature and make convenient to work in the field. The
flowers are immerged in the water for necessary period of time.

(C) Chemical treatment:

Certain chemical also known as gametocides when sprayed at the time of

flowering that induces male sterility in crop, means pollen become
temporarily non-functional. In this method care should be taken that only the
anthers are affected but not stigmas in the flower.

Gametocides: Certain chemical which is used to kill the pollen grain. e. g.

Name of Crop Chemical used
Cotton FW 450
Alfalfa 57 % Ethyl alcohol for 10 minutes
Wheat Ethyl-4-fluro oxinilate, Ethyl-4-chloro oxinilate,
Ethyl-2-Nitro oxinilate, Ethyl-1-2-Urethoxy oxinilate, etc.
Wheat, Rice, Spray of Ethrel
Sugarbeet

(D) Genetic emasculation:

In some crop plants, artificial emasculation is not required due to use of male
sterility which produces only sterile pollens. Genetic male sterility conditioned
by the presence of recessive genes. Cytoplasamic genetic male sterility is used to
produce commercial hybrid seed production in crops like maize, onion,
sorghum, bajra, etc. Similarly, self- incompatibility is also used for hybrid seed
production in diploid Brassicas.

30
 4) Bagging:

(i) To protect the emasculated flowers from undesirable pollens, it should

be covered with bag immediately.
(ii) Bag must be of proper kind and proper sized.

(iii) The bagged flowers are properly labeled.

5) Preparation of male flower:

(i) The bud of pollen parent should be enclosed in a bag before opening in order to
avoid contamination from the foreign pollen.

6) Crossing / Pollination:

1. The artificial transfer of pollens from desire male parent to the stigma of desire
emasculated flower of female parent is called crossing or pollination.
2. Pollination is done usually in the morning when the stigma is receptive.

The pollens are collected from the bagged flower of the male parent either in petri-dish or
in paper bag (maize, bajra, etc.) or direct male flower may be dusted on the stigma
(wheat, groundnut, brinjal, cotton, okra, etc.).

7) Bagging / Labeling:

 After pollination, the flowers are to be properly bagged and labeled.

8) Harvesting of cross seed:

1. The bags are removed and the cross seeds / hybrid seeds are harvested
and collected along with attached label.
2. After drying they are threshed.
3. Cross seeds are to be preserved properly to avoid stored pest damage

31
 EXPERIMENT: 6

HARDY-WEINBERG LAW

 WHAT IS HARDY- WEINBERG LAW?

 This law was proposed by two scientists Hardy (1908) in England and
Weinberg (1909) in Germany independently.
 This Law states that," in a large random mating population gene and
genotypic frequencies remain constant from generation to
generation in the absence of selection, mutation, migration or
random genetic drift."
 In a diploid species, two alleles A and a and there are three genotypes,
AA, Aa and aa.

 Now, if we denote the frequency of allele A as p and for allele a as q,

and sum of p and q is one i.e. p + q =1
 In the same way frequency of genotype

AA = p2 , Aa = 2pq and aa = q2, therefore

p2 (AA) + 2pq (Aa) + q2 (aa) = 1

When gene and genotypic frequencies remain constant, it will have said that
population is in equilibrium. This equilibrium is known as Hardy – Weinberg
equilibrium.

TERMINOLOGY:

 Random mating population: Cross pollinated crops are highly heterozygous

due to free intermating of plants within a population. They are often referred as
Random mating population or Panmictic population or Mendelian
Population.

32
 Mutation : A sudden heritable change in an individual and is generally due to a
structural change in a gene. Mutation may produce a new allele not present in the
population or may change the frequencies of existing alleles.

 Migration : The movement of individuals into a population from a different

population. Migration may have introduced new alleles into the population or may
change the frequencies of existing alleles.
 Random genetic drift : The change in gene and genotype frequencies of a
sample/ population entirely due to chance (small sample size, etc.).

 Selection : Differential reproduction rates of various genotypes are known as

selection.
 Gene frequency : The proportion of an allele A or a, in the population. OR The
proportion of gametes carrying an allele A or a is known as gene frequency.

 Genotypic frequency or zygotic frequency : The relative proportion of a

genotype AA, Aa or aa in the population is known as genotypic or zygotic
frequency.

Characteristics or properties of Random mating population:

1. Each genotype in the population is different in some degrees from others.

2. Each genotype is highly heterozygous.
3. Each genotype is largely out crossing or random breeding or cross pollinating.
4. Each genotype gives a variable progeny. Hence, gene and genotypic frequencies
are important in cross pollinated crops rather than genetic inheritance in self -
pollinated crops.

33
How to calculate gene and genotypic frequencies?

Suppose a sample of 100 individual drawn from a population which is made up

of AA = 25, Aa = 10 and aa = 65, hence N = 100,

Therefore, D = 25 H = 10 and R =65.

Now gene frequency of dominant A individual allele i.e. p

= [D + (½H)/N] = 25 + 5/100 = 0.3

Similarly, gene frequency for recessive allele a i.e. q

= [R + (½H)/N] = 65 + 5/100 = 0.7

Now, p + q = 1, therefore 0.3 +0.7 = 1.

Example – 1:

1) Which of the following population is in Hardy – Weinberg equilibrium?

Population Genotype frequencies
AA Aa aa
1 0.430 0.481 0.084
2 0.640 0.320 0.040
3 0.4225 0.4550 0.1225
4 0.0025 0.1970 0.8005
5 0.0081 0.0828 0.9091
2) What are the expected equilibrium frequencies for those above population
that are not in equilibrium?
3) How long will it take for this equilibrium value to be reached at random
population?

34
  Answer – 1 : Population –2.

Gene frequency of A = p = 0.64 + 0.16 =0.80

and Gene frequency of a = q = 0.04 + 0.16 =0.20

So, p + q = 1 = 0.80 + 0.20 = 1

 Here gene and genotype frequencies remain constant, so population -2 is in

Hardy–Weinberg equilibrium.
 Similarly, it is also calculated for all the remaining populations.

 Answer – 2: Expected genotypic equilibrium frequencies

AA = p2= (0.80) 2 Aa = 2pq = 2 x 0.8x0.2 and aa = q2 = (0.20)2

AA=0.64 Aa=0.32 and aa =0.04

 Answer – 3 : It will take only one generation to reach at random

population.

Example – 2: What is the frequency of heterozygotes (Aa) in a random mating

population, if the frequency of recessive phenotype (aa) is 0.09?

 Frequency of recessive phenotype (aa) i.e. q2 is 0.09,

Therefore, frequency of q= 0.3
So, the frequency of p =1-q =1-0.3=0.7

Genotype frequency of AA =p2 = (0.7)2 =0.49

Genotype frequency of Aa= 2pq= 2 x 0.7x0.3=0.42

Assignment:
1. What is Hardy Weinberg law? Explain it with a suitable example.

35
 EXPERIMENT: 7

ESTIMATION OF VARIABILITY PARAMETERS

VARIABILITY :

Variability means a difference among the individuals belonging to a single species

or different species within a population. The variability may be due to genotypes,
environments or due to the interaction of genotype x environment. There are two basic
types of variability. Genotypic variability and phenotypic variability.

1. Genotypic variability:

It is the component of variability which is due to genetic differences among the

individuals within a population. It is the main concern of the plant breeders.

2. Phenotypic variability:

It is the observable total variation present in a character of a population. It includes

both genotypic and environmental components of variability and as a result, its
magnitude differs under different environmental conditions. Thus, total variability
(phenotypic variability) can be partitioned into following components of variability as
under:

P=GXE
Where, P= Phenotypic variance
G = Genotypic variance
E = Environmental variance

A. Genotypic variability: Divided into three sub-component

1. Additive component: The component of variance arises by taking difference between

two homozygote is referred as additive variance.

2. Dominance component: The deviation of heterozygote from the average of the two
homozygote is referred as dominance variance. It is also called as Intra-allelic interaction.

36
 3. Epistatic component: The deviations arise as a consequence of inter-allelic interaction
is called epistatic variances. It also includes additive x additive, additive x dominance and
dominance x dominance interaction.

B. Environmental variability: It is due to environmental influence and is non-

heritable

Importance of Genetic Variability in Plant Breeding:

 The progress of any plant breeding programme for crop improvement depends
on the extent of the genotypic variability present in the base population.
 The assessment of the genetic variability present in the base population is the
pre requisite before the adoption of selection pressure in the variable
population and the efficiency of selection depends upon the identification of
genetic variability from the phenotypic expression of the character.
 The study of the genetic variability is necessary to determine the relative
importance of various plant characters with respect to yield in terms of genetic
variability.
 The parameters like range of variability, co efficient of variability, the
heritability and expected genetic advance are computed for the assessment of
genetic variability, and the gene action in expressing the genotypes which
reflects into effective selection in crop improvement programmes.
 Genetic variability for important agronomic traits in almost all the crops is
mainly due to the additive genetic variance. The non additive variance also
exists for many important traits are nearly all the crops but it is generally
smaller in magnitude than the additive component.
 Improvement in any crop plant needs sufficient genetic variability in the
experimental material which can be developed through hybridization,
mutation, polyploidy as well as by somaclonal variation.

37
  HERITABILITY:

 The ratio of genotypic variance to the phenotypic variance or total variance is

known as heritability. It is generally expressed in per cent. Thus, heritability
is the heritable portion of phenotypic variance.
 Heritability analyses estimate the relative contributions of differences in
genetic and non-genetic factors to the total phenotypic variance in a
population.
 It is a good index of the transmission of characters from parents to their
offspring.
 The estimates of heritability help the plant breeder in selection of elite
genotypes from diverse genetic populations.
 Heritability reflects the amount of variation in genotypic effects compared to
variation in environmental effects.
 Heritability can be made larger by diversifying the genetic background, e.g.,
by using only very outbreed individuals.
 On the other hand, smaller heritability can be generated by using inbred
individuals.
 Heritability does not quantify the extent to which genes and environment
actually determine a phenotype, let alone the extent to which changes in genes
and environment could change phenotypic values.

Types of Heritability:

Depending upon the components of variance used as numerator in the

calculation, heritability is of two types, viz., broad sense heritability and narrow
sense heritability.

i) Broad sense Heritability:

 It is the ratio of genotypic variance to total or phenotypic variance.

H2 = [Genotypic Variance / Phenotypic Variance] x100

38
 The features of broad sense heritability:
1. It can be estimated from both parental as well as segregating populations

2. It is estimated from total genetic variance

3. It is more useful in animal breeding than in plant breeding

4. It is useful in the selection of elite types from homozygous material

ii) Narrow sense heritability:

 It is the ratio of additive or fixable genetic variance to the total or

phenotypic variance.

h2 = [Additive Genetic Variance/Phenotypic Variance] x100

 The parameter h2 is the narrow sense heritability.

It plays an important role in the selection of elite genotypes from the segregating
populations.

The features of narrow sense heritability:

1. For estimation of narrow sense heritability, crosses have to be made in definite

fashion
2. It is estimated from additive genetic variance.

3. It is useful in both, plant breeding as well as animal breeding.

4. It is useful in the selection of elite types from segregating populations.

Difference between broad sense heritability and narrow sense heritability:

Broad sense heritability Narrow sense heritability

1. Estimated from total genetic Estimated from additive genetic
Variance variance

39
 2. Can be estimated from both Requires crossing in definite fashion
parental and segregating
Material
3. More useful in animal breeding Useful in both plant and animal
breeding
4. Useful in selection of elite types Useful in selection of elite types
from homozygous lines from segregating material

Estimation of Genetic Variability, Heritability and Genetic Advance :

The genetic variability present in breeding populations can be assessed by three ways

i. By using simple measures of variability

ii. By estimating the various components of variance and
iii. By studying the genetic diversity within a population

1). By using simple measure of variability:

When the experimental material is not replicated, the phenotypic (total) variability
present in breeding materials can be assessed by using following simple measures of
variability:

 Range: It is the difference between the highest and the lowest values present in
the observation of a sample.

Range = Maximum value – Minimum value

 Arithmetic Mean: It is also known as simple mean or average and it is computed

by dividing sum of all the observations in a sample by their number.

Arithmatic Mean (X) = ∑ X / n × 100

Where, ∑ X= summation of observation of all the samples

n = Number of observation in a sample

40
  Standard Deviation: It is the square root of variance and designated as SD (in
case of sample)

Standard = √S2
Deviation

 Coefficient of variation: It is an important measure of variability and is

commonly represented by CV. It is the percent ratio of standard deviation of a
sample to its mean.

Coefficient of variation % = SD/ X × 100

 Standard Error: It is measure of uncontrolled variation in a sample

Standard Error = SD/ √n × 100

2. By estimating the various components of variance:

The magnitude of phenotypic (total) variation present in a breeding population may be
portioned into several components of variation using appropriate experimental design in
replicated trials (discussed briefly in chapter-9).

3. By studying the genetic diversity within a population:

Two important biometrically techniques used for this purpose are as under:

1. Metroglyph analysis and index score method

2. D2statistics

Assignment:

1. Define/Explain the following

1 Variability 3 Standard Error

2 Heritability 4 Coefficient of variation

41
 EXPERIMENT: 8

FIELD EXPERIMENTATION

TERMINOLOGIES:

1. Treatment:

The objects of comparison which an experimenter has to try out in the field for
assessing their values are known as ‘Treatment.’e.g., varieties, manures,
cultivation practices, methods of seed treatments, insecticides, etc., are the
examples of treatments.

2. Experimental material:

The material on which the experiment is performed is called experimental

material. e.g. a set of varieties, a set of various sources of nitrogenous fertilizer,
a set of animals, etc.
 The experimental material is divided into a number of ultimate smaller units to
which the treatments are applied is known as ‘Experimental Units.’
 The experimental unit may be a plot of land, a patient in hospital, a cow, a
group of pigs in pen, a batch of seeds, etc

3. Uniformity trials:

A uniformity trial consists in growing in a field or piece of land a particular

crop with a uniform treatment, dividing the field into small units and harvesting
and recording the procedure from each of these units separately.

4. Experimental Error:

In addition to the uniformity of soil, there are other factors also which are
beyond the control of the experimenter and are responsible for the differences
in yield to a great extent even though there may be no real differences among

42
 the treatments. This variation due to uncontrolled factors known as
‘Experimental Error.’

Basic Principles of Field Experimentation:

There are three basic principles of field experimentation.

1) Replication 2) Randomization 3) Local Control

1) Replication:

 The repetition of the treatments under investigation is known as ‘Replication’.

 We know that the variation in soil fertility over the field is so great that all the
treatments do not get equal chance of experiencing every type of environment in
the field, if one treatment is allowed to one plot only.

 Some of the treatments which are applied to more fertile plots will be an
advantageous position as compared with those which are applied to less fertile
plots and are, biased. As a very simple illustration, we can take the case of an
experiment in which the two varieties for seed production are to be tested on a
piece of land. If the piece of land be divided into two equal plots and one variety
be sown in one plot and the other in the other, the difference in the yields of the
two varieties will not the real differential effect of the same, as they area
affected by varying soil fertility of the field also. It may be just possible that this
difference in the two yields may be due to the difference in soil fertility in the
two plots and not due to the difference in the varieties.

 If the land is divided into a number of smaller plots of equal size and in half of
the plots, one variety be sown and in the remaining half the other variety be
sown, selecting the plots for each variety by a random process, the two varieties
will be affected equally by the varying fertility of the land. Thus, the difference
between the yields of the two varieties would represent a valid estimate of the
differential effect of the two varieties.

43
 Advantages of Replications:
1. In order to obtain a greater precision in the field experiments, the most
effective method is to increases the number of replications. This will
reduce the error. The increase in precision can be done in two ways:

a. We know that the standard error of the mean is S / n where, ‘S’ is the
estimate of the standard deviation and ‘n’ is the number of observations on
which the mean is based. In this case n will represent the number of
replications. Now, it can be easily observed that the standard error of the
mean decreases as n increases and hence, the precision is increased. As the
smaller plots are more homogenous in fertility, it is better to keep the
experimental area the same and increases the number of replications by
shortening the plot size.

b. Another phase of increased precision arises from the distribution of t for

different degrees of freedom. As the number of degrees of freedom
increases, the value of t decreases with the result that the confidence
interval also decreases. The shortening of confidence interval is sufficient
proof of increased accuracy.

2. To estimate of error of the experiment for the purpose of comparisons.

The error of the experiment arises from the differences between the plots
of the same treatment. If, therefore, there is only one plot for each
treatment, there is no possibility of obtaining a measure of random soil
variability that can be used as an error in the test of significance. Without
the estimate of error, comparisons are not possible and without
replications estimate of error is not possible. Hence, replications are
absolutely essential for the experiment.

44
2) Randomization:

 The allocation of different treatments to the different experimental plots

by a random process is known as ‘Randomization of the treatments.’

 We also know that the comparison between the treatments will be valid

only when they are studied under equal environmental effects. In that case the
treatments get scope of showing their merits. This is possible by randomization
of the treatments.

3) Local Control:

 The principle of making use of greater homogeneity in groups of experimental

units (e.g. blocks of a number of plots homogeneous within themselves) for
reducing the experimental error, as explained above, is known as ‘Local
Control.’

 As a lower experimental error helps in detecting the smaller real differences

between the treatments, it is desirable that it should be reduced as far as
practically possible. This is possible by making use of the principle that the
adjacent areas are relatively more homogeneous than those widely separated.
In the field we find that the fertility of the field may be classified in two, viz.,
i) A major fertility variation which is usually marked by a fertility gradient.
ii) Sporadic or scattered fertility variations, which are not systematic but are
distributed in patches.

 Now, if we divide the whole field into blocks, which are homogeneous within
them, we can calculate the variance in yield due to the effects of the first type
of fertility variation and eliminate this variance for arriving at an estimate of
the variance due to chance error only.

 Randomizing the treatments within the blocks minimizes the effects of

45
 scattered fertility variations.

EXPERIMENTAL DESIGN:

 The choice of experimental design depends on the number and nature of the
treatments under study. It also depends on the object of the experiment.

The following designs are used under specific situations:

1. Completely Randomized Design (CRD): It is used when the

experimental material is limited and homogenous such as in the pot
experiments on soil, laboratory experiments, etc.

2. Randomized Block Design (RBD): It is used when the fertility

gradient in the field is in one direction. It may be adopted up to 20
treatments without an any appreciable loss of efficiency.

3. Latin Square Design (LSD):It is used when the fertility gradient is in

two directions. It may have adopted for number of treatments ranging
from 5 to 12 in most of the situations.

4. Factorial Experiment: When there are several factors with different

levels to be studied simultaneously with the same precision.

5. Split Plot Design (SPT): It is used when the factors are such that some
of them require larger plots (like irrigation, depth of ploughing, sowing
dates, etc) and some require smaller plots may be studied with different
precision.

6. Incomplete Block Design (IBD): It is used when the number of

treatments is sufficiently large.
Size and Shape of plots:

 There is no particular size or shape of the plots which may be said to be

the best for all circumstances.

46
  The plot size increases, coefficient of variation (CV) of plots decreases.
Therefore, it is better to increase the plot size up to the extent of 1/ 10 acre.
 If the plot size is greater than this, the increase in soil heterogeneity within
the plots affects any advantage obtained by increasing the plot size.
 However, an increase in the number of replications with reduced plot size
leads to a more precise treatment comparison, when the land and other
facilities are limited.
 So far as the shape of the plot is concerned it can be anything, square,
rectangle or a narrow long strip
 The dimensions of the plots are so chosen as to utilize the whole of the
experimental site effective and give a correct field layout.

Border Effect:

 The yield or any other character of the plants is affected in case of those plants
which are nearer to the borders of the plots with the result that the border plant
differ from the plants of the central portions of the plots with respect to the yield
or other character understudy.
 The growth of the plants is affected by the growth of a more vigorous variety of
the neighboring plots.
 The plants of the central plots remain unaffected by this growth.

 In manurial experiments, the manure from the manured plots might seep into the
un-manured plots and affect the plants upto some distance beyond the border.
Similarly the presence of the adjacent grass, drainage or unsown land also
affects the growth of the crop on the border of the plots.
 In order to ensure that yield or measurements of the character under study in
each plot has been only from the area which is unaffected by above mentioned
factors, non experimental border of sufficient width must be left around the plot.

47
 Assignment:

1. Define the following:

1. Treatment 2.Experimentalmaterial 3. Experimental Unit
4. Replication 5.Randomization 6. Local Control

2. Explain the basic principles of experimentation.

3. Write the advantages of replication.
4. Enlist the various experimental designs which can be used in different situations.
5. Under what circumstances will you used CRD, RBD, LSD, SPT and IBD?

48
 EXPERIMENT: 9

RANDOMIZED BLOCK DESIGN

 The magnitude of phenotypic (total) variation present in a breeding population

may be portioned into several components of variation using appropriate
experimental design in replicated trials.

 In randomized block design (RBD), the structure of analysis of analysis of

variance (ANOVA) is as follows:

Source d.f. S.S. M.S. Cal.F. Expected M.S.

Replication r-1 Rss MR MR /ME 2e+ g2r
Genotype g-1 Gss MG MG/ME 2e+ r2r
Error (r-1) (g-1) Ess ME --- 2e
Total rg-1 Tss -- ---

SEm = √Error M.S/r × 100

C.D. = SEm×√2 t0.0 5at error d.f.
C.V. % = √Error M.S/ X × 100

From the ANOVA of Randomized Block Design, the following components of variance
may be calculated:

1) Error variance (2e) = Me

2) Genotypic variance (2g) = (Mg – Me)/r
3) Phenotypic variance (2p) = 2g+ 2e
4) Genotypic coefficient = √2g/ X×100
of variation (GCV)
5) Phenotypic coefficient = √2p/ X ×100
of variation (PCV)

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 6) Heritability (h2b) = 2g/2p ×100
7) Genetic Advance (GA) = K ×p× H
Where, K=2.06 (selection differential)
8) GA as % of mean = G.A./ X ×100

Relative importance of Variability parameters:

1. Genotypic variance: Genotypic variance only measures the amount of

genetic variability present in a character of a population. However, it
does not provide a measure to compare the genetic variability among
various characters of a population

2. GCV: Genotypic coefficient of variation measures the range of genetic

variability present in a character of a population and also provides a
measure to compare the genetic variability among various plant
characters within and between different populations. However, it does
not asses the amount of heritable variation in relation to total variation
present in a character of a population.

3. Heritability: Heritability in broad sense provides measure to assess the

amount of heritable variation in relation to total variation present in a
character of a population and also to compare the heritability among
various plant characters. It only indicates the effectiveness with which
selection of genotypes can be based on the phenotypic performance.
However, it fails to indicate the expected genetic progress of selection.

4. Genetic advance: Expected genetic advance provides indication of the

genetic progress which can be expected from selection of superior
genotypes from the population, but fails to compare the expected genetic
progress of selection among various plant characters.

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 5. Genetic Advance (%): Expected genetic advance as % of mean provides a
measure to compare the expected genetic progress of selection among various
plant characters within a population and between populations.

Example

Eight varieties of barley were tested in RBD with four replication and the
observations were recorded on number of ears per plant. Calculate variability
parameters, heritability and genetic advance.

Character: Number of ears per plant

Varieties R-I R-II R-III R-IV TOTAL

1 50.2 41.4 36.2 39.8 167.6
2 41.8 47.2 39.6 46.6 175.2
3 39.2 37.6 38.8 33.6 149.2
4 37.8 49.6 35.4 41.8 164.6
5 35.6 31.4 33.2 29.8 130.0
6 53.4 50.2 49.6 57.8 211.0
7 43.8 46.8 41.4 43.6 175.6
8 50.6 47.8 41.8 46.8 187.0
Total 352.4 352.0 316.0 339.8 1360.2

Analysis of variance: Total S.S

Mean (X) = G.T. = 1360.2

32 = 42.506
N
Correction = (G.T.)2 = (1360.2)2
factor (C.F.) = 57817.00
N 32

Total S.S. = [(50.2)2 + (41.8)2 + …… = 1422.959

+ (46.8)2] - C.F.

Treatment = (167.6)2 + (175.2)2 + =

1023.989
S.S. ….. + (187.0)2/4 - C.F.

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Replication = (352.4)2 + (352.0)2 + = 109.224
S.S. (316.0)2 + (339.8)2/8- C.F.

Error S.S. = Total S.S.- Replication = 1422.959 – 109.224 = 289.746

S.S. - Treatment S.S. – 1023.989

ANOVA

Source d.f. S.S. M.S. Cal.F.

Replication 3 109.224 36.408 2.639 (NS)
Genotype 7 1023.989 146.284(M1) 10.602**
Error 21 289.746 13.797(M2) --
Total 31 1422.959 ---- ---

Calculation of variability parameters:

SEm = √Error M.S./r × 100
= √13.797/4 = 1.857

C.D. = S.Em× 2 × t 0.05 at error d.f.

= 1.857 × 1.414 × 2.080 = 5.46

C.V. % = √Error M.S./ X × 100

= √13.797/42.506 × 100 = 8.7386

From the ANOVA of Randomized Block Design, the following components of variance
may be calculated:
1) Error variance (2e) = Me
= 13.797

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2) Genotypic variance (2g) = (Mg – Me)/r
= (146.284 – 13.797)/4 = 33.122

3) Phenotypic variance (2p) = 2g + 2e

=  = 

4) Genotypic coefficient = √2g/ X ×100
of variation (GCV)
= √33.122/42.506× 100 = 13.5397

5) Phenotypic coefficient = √2p/ X ×100

of variation (PCV)
= √46.919/42.506 × 100 = 16.1147

6) Heritability (h2b) = 2g/2p ×100

= × = 

7) Genetic Advance (GA) = K ×p× H
= 2.06×6.849×0.7059 = 9.95

8) GA as % of mean = G.A./ X ×100

= 9.95/42.506× 100 = 23.43

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 EXPERIMENT: 10

COMBINING ABILITY
Combining ability:
 The concept of general and specific combining ability as a measure of
gene action was first proposed by Sprague and Tatum (1942).

 Combining ability is the relative ability of a strain to transmit desirable

performance to progeny upon hybridization with other strains.

There are two types of combining ability:

1) General combining ability (GCA):

 It is the average performance of a genetic strain in a series of cross

combinations, estimated from the performance of F1’s from crosses. OR
 It is the average ability of a parent to combine with a set of other parents in
hybridization.

 Characteristics of General combining ability (GCA):

 GCA is related to parents only.

 GCA effect of a parent (P) is calculated as (G –M)

 where, G=Mean value of crosses in which P is a common parent, and

M = General mean

 Progeny produced by GCA is half-sibs (H.S.)

 GCA variance is attributed to additive genetic variance and additive x

additive interaction variance which are considered as fixable.
 The knowledge of GCA helps to evaluate the parents of broad genetic
base.

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2) Specific combining ability (SCA):

It is the deviation in the performance of a specific cross combination from the

performance predicted on the basis of general combining ability of the parents
involved in the cross. OR

It is used to designate those cases in which certain combinations do relatively

better or worse than that would be expected on the basis of average performance
of the parents involved in the cross.

 Characteristics of Specific combining ability (SCA):

 SCA is related to crosses (hybrids) only.

 The knowledge of SCA helps to identify the hybrids.
 High SCA may be due to good x good, good x poor or poor x poor combiners.
 If high SCA is due to high x high GCA effects, one can exploit the hybrid vigor
in F1.
 If high SCA is due to high x low GCA effects, such parents are used for
development of sister lines which are used for the development of synthetic
varieties. Based on this knowledge, further breeding programme is to be
formulated.
 If high SCA is due to low x low GCA effects, there are least chance of
improving such parents.
 SCA effect of a cross is calculated as (S – G1 – G2 –M)
 Where, S = Mean value of a specific crosses,
 G1& G2 = GCA effects of two parents involved in a cross, and
M = General mean.
 Progeny produced by SCA is full-sibs (F.S.)
 SCA variance is attributed to dominance genetic variance and additive x
dominance and dominance x dominance interaction variance which are

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 considered as Non-Fixable.
 Kempthorne (1957) defined GCA and SCA variances in terms of covariance of
half and full sibs in a random mating population:

Methods to estimate Combining ability:

1. Inbred – variety cross / top cross (Jenkins & Brunson,1932)

2. Polycross (Tysdal et al.,1942)

3. Diallel cross analysis (Griffing, 1956 a &b)

4. Line x Tester analysis (Kempthorne,1957)

Utility of combining ability in Plant Breeding:

1. It helps to identify the parents having good potential to transmit desirable

characteristics to their progenies

2. It helps to sort out the promising crosses for yield and its components.

3. It also provides the information regarding the nature and magnitude of

gene action involved in the inheritance of traits.

4. It is valuable for adoption of efficient breeding programme.

Selection of parents based on GCA is more desirable as compared to that

based on per se performance. Why?

 Parents are selected on the basis of per se performance such parents may be good
but transmission ability to their progenies may or may not be good. While
parents selected on the basis of GCA denotes good transmission ability to their
progenies.

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  In addition nicking ability of parents is also based on GCA. Therefore, parents
selected on the basis of GCA are more desirable as compared to selection of
parents on the basis of per se performance.

How combining ability is superior over simple hybridization between any two
parents at a time?

 The knowledge of combining ability of parents helps to predict the

performance of resulting hybrids and accordingly one can formulate the
breeding programme, while in case of simple hybridization between any two
parents; one cannot formulate the breeding programme.

Relationship between Combining ability and Gene Action:

1). If only GCA variance is significant:

Additive gene action is important. In such cases, pedigree method of

selection may be effective and that should be followed.
2). If only SCA variance is significant:
Non-additive gene action is important. In such cases, exploitation of heterosis in
F1 should be followed.
3). If both GCA and SCA variances are significant:
Both additive and non-additive gene actions are important. In such cases,
biparental mating approach or reciprocal recurrent selection by alternate selfing
and inter mating should be followed which helps in isolation of desirable
transgressive segregants in segregating generations

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 TYPES OF CROSSES

1) SINGLE CROSS:
 It was proposed by G.H. Shull (1909).
 A cross between two pure lines or inbred lines. e.g.

AxB F1 (AB)

 The number of possible single crosses produces from various pure lines or
inbred lines can be calculated as follows:

 Number of possible single cross without reciprocal = [n ( n-1 )] /2

 Number of possible single cross with reciprocal = [n ( n-1)]
Where, n = Number of pure lines or inbred lines.

 Uses:
i) To study the combining ability of different pure lines or inbred lines.
ii) To predict the performance of double cross combination.

2) DOUBLE CROSS:

 It was proposed by D.F. Jones (1918).

 A cross between two F1 or hybrids. e.g.

A x B C x D

F1 x F1

F1 (Double cross)

 The number of possible double crosses produces from various pure

lines or inbred lines can be calculated as follows:

 Number of possible double cross = [n ( n-1 ) ( n-2 ) ( n-3 )] /8

Where, n = Number of pure lines or inbred lines.

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  Uses:
i) It is used to produce double cross hybrid for commercial planting.
e.g. Double cross hybrids are available in maize.

ii) To study the combining ability of the parents.

3) THREE WAY CROSS:

 A cross is made between the F1 of a single cross and an inbred line.

 F1 is always used as female and inbred line as male.e.g.

AxB

F1 x C

4) MULTIPLE CROSS or Complex cross or Convergent cross :

F1 (Three way cross)
 When more than four parents are involved in a cross, it is known as multiple
cross or complex cross or convergent cross.
 It was first introduced by Harlan et al., (1940) in barley.
 The concept is to combine desirable traits from various parents in a systemic
series of crossing cycle.
 After the first parental single cross combination is obtained, the subsequent
crosses are to be made in a pyramid fashion.
 For example

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 AxB CxD E xF GxH 1st crossing cycle

AB x CD EF x GH 2nd crossing cycle

3rd
ABCDEFGH Multiple cross

ABCD x EFGH crossing cycle

 Uses:
i) To create more amount of genetic variability for different characters.
ii) To combine different desirable genes which are scattered into different parents.

 Disadvantages:
i) It is time and labour consuming method.
ii) Undesirable recombination is also brought together.
iii) The probability of obtaining the plants with all desirable genes is decreasing
with increasing in number of parental lines.

5) BACK CROSS:
 A cross between hybrid (F1) and one of its parents. e.g.

P1 (AA) ×P2 (aa) P1 (AA) ×P2 (aa)

F1 ×P1 (AA) F1 ×P2 (aa)

BC1 BC2

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  Uses:
i) To transfer a desirable character from the donor parent to the recipient
parent this is lacking in recipient parent.

ii) To produce a multiline varieties.

6) TEST CROSS:
 A cross of hybrid (F1) with a homozygous recessive parent. e.g

P1 (AA) × P2 (aa)

F1 x P2 (aa) homozygous recessive parent

Test cross progeny

 Uses:
i) For determining the kind and type of gamete produced by F1.
ii) For linkage studies.

7) TOP CROSS OR INBRED VARIETY CROSS:

 The cross made between an inbred and open pollinated variety (OPV).
 It was proposed by Davis (1929).
 Open pollinated variety is used as a tester for studying the combining
ability of inbred line.

Inbred line x Open Pollinated Variety

8) DIALLEL CROSS:

 Crossing among the genotypes in all possible combinations.

 It is mainly used to study the general combining ability and specific

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 combining ability of parents and hybrid, respectively.

 General Combining Ability (GCA) : An average performance of a strain in a

series of crosses estimated from the performance of F1 hybrids.

 Specific Combining Ability (SCA) : It is the deviation from the performance of

a cross predicted on the basis of GCA of the parents involved in a cross.

Diallel mating scheme for seven parents viz., A, B,C, D, E, F and G (Half diallel)
AxB BxC CxD DxE ExF FxG
AxC BxD CxE DxF ExG
AxD BxE CxF DxG
AxE BxF CxG
AxF BxG
AxG

9) POLY CROSS:

 The open pollination of a group of genotypes in isolation from other compatible

genotypes in such a way to promote random mating. In this, all the genotypes
have equal chances of pollination.

 It was proposed by Tysdal, Kiessel bach and Wastover (1942).

 It is generally used in cross pollinated crops especially in grasses and forage
crops in which hand pollination is very difficult due to very small and tiny size
of flowers.

Assignment:
1. Explain different types of crosses with example and write its uses.
2. What is combining ability? Write its types and compare it.
3. Write characteristics of GCA and SCA.
4. Give the utility of combining ability in plant breeding.

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5. Write the relationship between combining ability and gene action.
6. Justify: Selection of parents based on GCA is more desirable than that based on
per se performance.

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