Hybridization: Techniques
and Consequences
Hybridization
The mating or crossing of two plants or lines of dissimilar
genotype is called hybridization.
Natural variability present in Self-pollinated populations is
exhausted quickly when they are subjected to selection.
Therefore for further improvement new genetic variability
has to be created which is easily and most commonly
achieved by crossing two different purelines
Genotype peculiarities of two or more different varieties or
species of plants could be brought together only by crossing
them.
Hybridization….
In plant hybridization, one of the plants is taken as the
female plant and the other as the male plant.
Pollen grains from the male parent are made to
pollinate the stigma of the flowers of the female
parent.
The seeds obtained from such a cross are called F1
seeds and the progeny raised from it is called F1 (First
Filial) generation.
The F1 is selfed to produce F2 and the subsequent
generations like F3, F4 etc are raised in the same
way(selfing).
These generations are called segregating generations
and they are handled differently based on the scope
and objectives of the breeding program.
OBJECTIVES OF HYBRIDIZATION
The major objectives of hybridization are
A. To artificially create a variable population for the
selection of types with desired combination of
characters.
B. Combination Breeding
Combination breeding is the transfer of one or more
characters from other varieties to a Single genotypes
These characters may be oligogenic or polygenic.
In this way, genes for disease resistance, quality traits etc
can be transferred.
C. Transgressive Breeding
Transgressive breeding is based on
transgressive variation of characters in
segregating generations like F2.
A cross is made between two strains of plants
and the F2 is screened for transgressive
variations.
Transgressive segregation is the segregation
of characters beyond the parental limits, in the
segregating generations like F2.
D. Production of Hybrids
Hybrid (F1) plants show higher vigor and
yield when compared to parents, in some
cases.
This phenomenon is called hybrid vigor and
heterosis
F1 seeds can be raised in bulk through
hybridization and distributed directly for
cultivation, especially in cross-pollinating
crops.
TYPES OF HYBRIDIZATION
Based on the genetic difference between
parents, hybridization can be classified into:
1.Inter-varietal Hybridization
2.Distant Hybridization
Inter-varietal/Intra specific
hybridization
The cross between the members of the same species (intra-
specific) is called inter-varietal hybridization.
It is most commonly used in crop improvement programs
In this type of hybridization, different cross patterns can be
used.
Inter varietal crosses are called simple or complex depending on
the number of parents involved
1. Simple Cross
2. Complex Crosses
3. Diallel Cross, Half diallel cross:
4. Partial diallel cross.
Simple Cross
• In this case, two parents are used to produce an F1
hybrid. Such cross is also called single cross
Parent A X Parent B.
SC F1
Single Cross
Complex crosses
• It involves more than two parents are.
• Such crosses are also known as convergent crosses
since they bring genes from different sources together
to a single hybrid
It may be :
1. Three way cross:
2. Double Cross
3. More complex crosses
1. Three way cross
• Parent A X Parent B
SC (F1) Hybrid X Parent C
TWC Hybrid denoted by A/B//C
Fig: Three way cross hybrid
Double Cross
• Parent A X Parent B Parent C X Parent D
SC1 (F1) Hybrid X SC2(F1 hybrid)
DC Hybrid Cross is denoted by A/B//C/D
Fig: Double cross hybrid
Complex Crosses:
It includes usually morethan 4 parents
AXB CXD EXF GXH
SC 1 X SC2 SC3 X SC4
DC1 X DC2
Complex cross Hybrid
Diallel cross:
• Crossing a number of genotypes in all possible combinations
among a group of inbreed lines. It could be used to estimate of
both GCA and SCA as well as determination of effect of
reciprocal crosses. Total no. of crosses = n(n-1)
Half diallel cross : Crossing a number of genotpes in all possible
combination but no reciprocal of crosses are made.
No of crosses = n (n-1)/2
Partial diallel cross:
A diallel cross based on a specified sample of crosses from among
all possible crosses ( diallel scheme) among a number of
parental lines.
It is defined as the no of sample crosses per parent or per array in
all possible combination of a given set of parents.
Total no. of sample cross = ns/2, n= no of parent, s = no. of sample
cross /plant
Distant hybridization
• Hybridization includes crosses between different species of the
same genus or of different genera.
• It is hybridization beyond species level
• Thus, it may be interspecific (intra-generic) or inter-generic.
Generally, the objective of such crosses is to transfer one or few
simply inherited characters like disease resistant
E.g Gene for resistance against various pathotypes of Bacterial
blight (Xanthomonas oryzae pv oryzae) have been transfer from
wild rice Oryza rufipogon and O. nivara
Distant hybridization is likely to become increasingly important in
the correction of specific defects of crop species.
In many cases wild species contribute valuable quality genes and
even yield genes
Hybridization procedure:
Advance planning:
• Development of a variety takes 7-8 years
• Breeder has to foresee the requirements about
8-9 years in advance
• He/she should be familiar with requirement of
market such as quality, disease, pest, abiotic
resistance, increased yield etc.
• Further he/she should clearly define what type
of variety is to be produce
The steps involving hybridization are as:
Choice of parents:
• It depends on the objective of breeding programme.
• Increase yield is the common objective of the breeder
so at least one of the parents involved in a cross should
be a well-adapted and proven variety in the target
area for which the new variety is being developed.
• Whereas the other variety should be having the
characters that are weak or absent in the adapted
variety/lines.
• Parents with good combining ability should selected.
• Make sure all characters sought to be improved are
present in one or the other parent.
• 2, 3 or more parents can be selected depending upon
necessity
Evaluation of parents:
• If the performance of parents in the area where
breeding is to be done is known, evaluation is
not necessary. Otherwise it should be done.
• Screening of for disease reaction for introduced
parents should be done.
• Selfing of parents is performed to eliminate the
undesirable characters and obtain inbreeds.
Planting/sowing
• Normally different genotypes have different time of anthesis.
• Planting of genotypes should be done in such a way that time of anthesis
of female parent should coincide with shedding of viable pollen in male
parent.
• Planting should thinner than normal planting.
• Greenhouse planting in small pot will be good.
• Parent 1 Planting date Anthesis date
• Parent 2 planting date Anthesis date
Emasculation:
• Removal of stamens or anthers or killing of
pollen grains of a flower without affecting the
female reproductive organs is known as
emasculation.
• Purpose of emasculation is to prevent self
fertilization.
• In dioecious species male plants are removed
and in monoecious species male flower is
removed but in bisexual flower emasculation
is essential
Different methods of emasculation
Hand emasculation:
• In species with relatively
large flowers, stamens or
anthers are removed by the
help of forceps .
• Care should be taken to
remove all the anthers from
the flowers without breaking
them, without damage the
stigma and ovary.
• Should be done before the
anthers mature and the stigma
has become receptive.
Suction method:
• Useful in species with small flowers.
• Petals are removed with forceps exposing
anthers and stigma.
• Thin rubber or glass tube attached to a
suction hose is used to remove the anthers
• Considerable amount of self-pollination (up to
15%) is likely to occur
Hot/cold water or alcohol treatment:
• Pollen grains are more sensitive than the female
reproductive organs to both genetic and
environmental disturbances, so this property is
utilized to kill pollen grain with hot/cold/ water
treatment without damaging female reproductive
organs.
• Treatment should given before flower opens and
anther dehisce.
• For rice, 10 min treatment with hot water at 40 – 44
degree centigrade is adequate.
• With cold water at 0-60 C kills pollen grains
• Cold water is less effective than hot water
Alcohol treatment
• Rarely used method consists of immersing the
flower or inflorescence in alcohol of suitable
concentration followed by rinsing it with
water.
• Duration of treatment and concentration is
important
• E.g. 57 % alcohol treatment in sweet clover
for 10 seconds.
Genetic emasculation:
• Male sterility or self in-compatibility
techniques can be used to eliminate the
necessity of emasculation.
Chemical emasculation:
Many chemicals like sodium methyl arsenate
(MG2), ethephon, GA3, hybrex etc. induce
artificial non genetic male sterility and these
chemicals are called chemical hybridizing
agents(CHAs).
Pollination without emasculation
• Some crops have protogynous flowers and stigmas of
their flowers become receptive one or two days before
anthesis.
• In such crops , flowers buds that would open the next
day may be selected and directly pollinated without
emasculation because their anther will not yet be fully
mature.
• Use full in species like soybean that have small
flowers prone to damage during emasculation.
• In soyabean this method gave around 39 % pod set
as compared to about 2%-3% pod set with hand
emasculation.
Bagging:
Immediately after emasculation, the flowers or
inflorescence are enclosed in suitable bags of
appropriate size.
The main objectives of bagging are:
• To prevent contamination in male flowers and
• To prevent cross pollination in female flowers
The moisture and temperature are generally higher inside
the bags as compared to the outside.
Therefore bagging may promote fungus development on
the fruit or the spike. Removing the bags usually 2-3
days after pollination may prevent this.
Pollination:
• Pollination is the transfer of pollen grains from
anthers to the receptive stigmas of emasculated
flowers
• Preferably done in the morning hours(9-
11am).
• The pollen grains are collected from the
desired parent and kept on the stigmas of the
desired female parent.
Tagging/labelling:
The emasculated flowers are tagged just after
bagging.
Tags are available in different sizes.
Information like
• date of emasculation,
• date of pollination,
• name of the female parent,
• name of the male parent
• and the name of the breeder
is recorded on the tag with marker.
Harvesting and storing the F1 seeds:
• Harvest, thresh, properly dry and stored to
protect them from storage pest.
• Care should be taken to avoid contamination
of hybrid seed with other seeds.
• Seed of each crosses should be kept separately
with original tags to avoid confusion.
Raising of F1 hybrid
Larger F1 size, greater opportunity for rare
recombinations to occur but tedious to manage.
But wherever possible a larger F1 population is
more desirable than smaller one
F1 seed is sown carefully so that germination and
growth is successful.
F1 is usually allowed self pollination but in
backcross programme it is back crossed to one of
the parents.
Handling the F2 and later generation
• Selection of desirable plant type has to be done
in the segregating generations.
• Selection for qualitative traits is simple quick
but for quantitative traits is often difficult and
time consuming.
• Plants are selected on the basis of breeding
objectives following proper breeding
methods.
Consequences of Hybridization
• Production of new genotypes : segregation and
recombination of genes.
• The number of phenotypes produced rapidly with the
number of segregating genes.
• In F2 vast majority of plants would be heterozygous which
reduces effectiveness of selection.
• Linkage would decrease frequency of recombinant types
and increase the frequency of completely homozygous plants.
• Linkage between favourable genes make selection easier
while between unfavourable genes will be problem.
• In case of quantitative traits some recombination produce
superior genotypes which surpass both parents. These
recombinants are known as transgressive segregants
For n number of segregating genes
• Different types of gametes produced by F1=2n
• No. of different genotypes in F2 = 3n
• Smallest perfect F2 population size = 4n
• Number of phenotypes in F2
a. Full dominance = 2n
b. No dominance = 3n
Number of completely homozygous genotypes = 2n
The frequency of completely homozygous plants in
any segregating generations is give by
= {(2m-1)/2m}n where, m= no. of generations of self
pollination
