role of mutation breeding in crop improvement

1. Department of Genetics & Plant Breeding
Sardar Vallabhbhai Patel University of Agriculture and Technology
Meerut 250110
MASTER SEMINAR
Crop Improvement through Mutation
Breeding
Presented by
Mahesh Upadhyay
PG/A- 4839/19
M.Sc. (Ag.)
Course Co-ordinator
Dr. Pooran Chand
Professor
Course co cordinator
Dr. Atar Singh
Assistant Professor

2. 2
Introduction
Mutation and it’s types
Mutagenesis
Mutation breeding
Cos and Pros of mutation breeding
Way forward
Case Studies
Contents

3.  This change in the character of the individual
is due to change in number or sequence of
nucleotides.
 The term mutation was first used by Hugo de
Vries (1900) to describe the sudden
phenotypic changes which were heritable,
while working with Oenothera lamarckiana.
Mutation and its Historical background
 Mutation can be defined as a sudden heritable change in the
character of an organism which is not due to either segregation or
recombination.
Hugo de Vries

4.  English farmer Seth Wright recorded case of mutation first time in
1791 in male lamb with unusual short legs.
 Systematic study of mutation was started during 1910 when Morgan
genetically analyzed white eye mutant of Drosophila.
 H. J. Muller induced mutation in Drosophila by using X- rays in 1927
; he was awarded with Nobel prize in 1946.
 C. Auerbach and J.N. Robson in 1946 used chemicals to induce
mutations in Drosophila.
 The first plant breeding programme using mutations (mutation
breeding) was initiated in 1929 in Sweden, Germany and Russia.
 The term mutation breeding was first coined by Freisleben and Lein
(1944).
 Father of mutation breeding - Ake Gustafsson
HISTORY

5.  Muton: The smallest unit of gene capable of undergoing mutation and it
is represented by a nucleotide.
 Mutator Gene: A gene which causes another gene or genes to undergo
spontaneous mutation.
 Mutable Genes: Genes which show very high rates of mutation as
compared to other genes.
 Mutant: An organism or cell showing a mutant phenotype due to mutant
allele of a gene.
 Mutagen: A physical or chemical agent which induces mutation.
 Hot Spots: Highly mutable sites with in a gene.
 Gene mutations or Point mutations: The changes which alter the
chemical structure of a gene at molecular level.
Important terms related to Mutation

6. Classification Of Mutation

7. Based on the survival of an individual:
 Lethal mutation – when mutation causes death of all individuals
undergoing mutation are known as lethal.
 Sub lethal mutation - causes death of 90% individuals.
 Sub vital mutation– such mutation kills less than 90%
individuals.
 Vital mutation -when mutation don’t affect the survival of an
individual are known as vital mutation.
 Supervital mutation – This kind of mutation enhances the
survival of individual.

8. 1. Spontaneous mutation
 Spontaneous mutation occurs naturally without any cause. The rate of
spontaneous mutation is very slow.
 Rate of spontaneous mutation is higher in eukaryotes than prokaryotes.
E.g. UV light of sunlight causing mutation in bacteria
2. Induced Mutation
 Mutations produced due to treatment with either a chemical or physical
agent are called induced mutation .
 The use of induced mutation for crop improvement program is known
as mutation breeding.
E.g. X- rays causing mutation in cereals
Based on causes of mutation

9. Based on tissue of origin:
1. Somatic mutation
 A mutation occurring in somatic cell is called somatic mutation.
 In asexually reproducing species somatic mutations transmits
progeny to the next progeny.
2. Germinal Mutation
 When mutation occur in gametic cells or reproductive cells are
known as germinal mutation.
 In sexually reproductive species only germinal mutation are
transmitted to the next generation.

10. Based on direction of mutation:
1. Forward mutation
 When mutation occurs from the normal/wild type allele to
mutant allele are known as forward mutation.
2. Reverse mutation
 When mutation occurs in reverse direction from mutant allele
to the normal/wild type allele are known as reverse mutation.

11. Type of trait affected:
1. Visible mutation- Those mutations which affect on phenotype
and can be detected by normal observations are known as
visible mutations.
2. Biochemical mutation- mutation which affect the production
of biochemicals and which does not show any phenotypic
character are known as biochemical mutation.

12. Characteristic features of mutation
 Mutations are mostly recessive and very rarely dominant.
 Most mutations have harmful effects and very few (less than 0.1 %) are
beneficial.
 Mutations may be due to a change in a gene, a group of genes or in
entire chromosome.
 Chromosomal mutations are generally lethal and such mutants do not
survive.
 If mutation occur at both loci simultaneously, the mutants can be
identified in M1 generation. However, if it is restricted to one locus
only the effect can be seen only in M2 generation.
Contd..

13.  Macro-mutations are visible and can be easily identified, while
micro mutations can not be seen with naked eye.
 Most mutants are of negative selection value.
 Mutation for altogether new character generally does not
occur.
 Mutations are random i.e. they can occur in any tissue or cell
of an organism. However some genes show higher mutation
rate than others.
 Mutations are recurrent i.e. the same mutation may occur
again and again.
 Induced mutations commonly show pleiotropy often due
mutation in closely linked genes.

14.  Occur naturally without any apparent cause.
 There are two possible sources of origin of these mutations.
1. Due to error during DNA replication.
2. Due to mutagenic effect of natural environment .
E.g : UV rays from sunlight
 The rate of spontaneous mutations is very low. 10-6
 Some varieties were developed through spontaneous mutations.
Mutations useful in case of Crop Improvement
1.Spontaneous mutations
S. No Crop Variety
1 Rice GEB-24, Dee-Geo-Woo-Gen
2 Wheat Norin
3 Groundnut TMV-10
4 Sorghum Co-4 (Coimbatore 4)

15.  Mutations can be induced artificially through treatment with either
physical or chemical mutagens. The rate of induced mutations is very high.
 EMS-induced mutation is a highly effective method and, therefore,
commonly used in crop breeding to develop improved crop varieties
2. Induced Mutation
S. No Crop Mutant Variety Original Variety Mutagens
1. Rice Jagannath T-141 X-rays
Mahasuri mutant Mahasuri γ-rays
2. Wheat Sarbati sonara Sonara-64 UV rays
NP-836 NP-799 X-rays
3. Sesame Kalika BM-3-7 EMS
4. Castor Aruna HC-6 Thermal neutrons
5. Cotton MCU-7 1143EE X-rays

16. Mutagenesis
Mutagenesis is defined as the change in the genetic information of an organism
in a stable manner by the use of physical and chemical mutagens.
Types of Mutagenesis
1. Random mutagenesis 2. Site-directed mutagenesis
 Random mutagenesis-
In this an organism is exposed to a physical or chemical mutagen, mutations
are induced randomly in all genes of the organism. The desired mutant is selected
from the mutagenised population.
 Site-directed mutagenesis-
also called site-specific mutagenesis or oligonucleotide-directed mutagenesis,
in this a mutation is created at a defined site in a DNA molecule.

17. The agents which causes mutations is known as mutagen.
Types of mutagens:
 Physical mutagens
 Chemical mutagens
Mutagens
A. Physical mutagens
Include various types of radiations, viz., X-rays, γ-rays, α-rays, ß-
rays, fast neutrons, thermal or slow neutrons, UV rays etc.
The physical mutagens are classified into-
 Ionizing radiations: They work through the release of ions. They
have deep penetrating capacity. E.g : x-rays, γ-rays, α-particles etc.
 Non-ionizing radiations : They function through excitation and have
a very low penetrating capacity. E.g : UV rays

19. The primary research centres and institutes in India that
participated in the development and release of various
mutants:
 Indian Agricultural Research Institute (IARI)- New
Delhi
 Bhabha Atomic Research Centre- Mumbai
 Tamil Nadu Agricultural University -TN and
 National Botanical Research Institute - Lucknow, UP

20. PROCEDURE
FOR
MUTATION BREEDING

21. Choice of material
1
Choice of mutagen
2 Dose of the mutagen
3
Plant part to be
treated4
Giving mutagen treatment 5
Handling of the
mutagen-treated
population6
Validation7
PROCEDURE FOR MUTATION BREEDING

22.  Choice of material : It should be the best variety available in
crop and seed should be pure.
 Choice of mutagen : Generally chemical mutagens are more
preferred for seed treatment and radiations for the treatment of
vegetative parts.
 Dose of mutagen:
 Optimum mutagen dose is one, which produces maximum
frequency of mutations and causes the minimum killing.
 Close to LD50 dose is optimum. LD50 is the dose of
mutagen that kills 50% of the treated individuals.
Eg:- EMS – 0.3-1.5 %, for 2-6 hours

23. Plant
parts
Pollen
grains
Corms
Bulbs
Vegatative
propagules
Complete
plant
Seeds
Part of the Plant to be Treated

24. Handling of segregating population
M1 generation
 Seeds treated with chemical mutagens should be washed
thoroughly and be planted as soon as possible.
 Large M1 generation is raised from treated seeds (Wider
spacing).
Eg: 25,000 plants are to be grown to obtain a useful
mutation in M1 generation.
 Mutagens with high mutation frequency - M1 generation
size can be reduced.
Contd…

25.  The M1 plants should not be allowed to cross pollinate.
 M1 population should be planted 75-100 m apart from the
parental or other genotypes of the same crop species.
 Mechanical isolation.
 M1 generation Dominant mutations are selected each plant
selfed and harvested separately for M2.

28. TILLING and ECO-
TILLING4
3
Microarray design probe
2
Genome-wide chips snp
1
Methods for Validation of mutants
PCR screening design

29. TILLING (Targeting Induced Local Lesions IN Genomes) and
ECOTILLING are methods used in detecting induced or naturally
occurring mutations in many species.
ECOTILLING Technique use to survey natural variation in genes .
TILLING and ECOTILLING have recently been used for the
detection of both induced mutation and natural DNA polymorphism.
The chemical mutagen EMS generates mostly SNPs, and can be
controlled to produce a high density of point mutations, causing a
variety of lesions including nonsense and missense mutations.
(McCallum et al.,2000)
TILLING AND ECOTILLING

30.  DNA is collected from a mutagenized population (TILLING), or a
natural population (EcoTILLING). For TILLING, DNAs from up
to eight individuals are pooled.
 Typical EcoTILLING assays do not use sample pooling, but
pooling has been used to discover rare natural single-nucleotide
changes.
 After extraction and pooling, samples are typically arrayed into a
96-well format.
 The target region is amplified by PCR with gene-specific primers
that are end-labeled with fluorescent dyes.
 Following PCR, samples are denatured and annealed to form
heteroduplexes that become the substrate for enzymatic mismatch
cleavage.
 Cleaved bands representing mutations or polymorphisms are
visualized using denaturing polyacrylamide gel electrophoresis.
Procedure

31. The Basic Steps For Typical TILLING And Ecotilling Assays
(Till et al., 2006)

32. Gamma Gardens
are a form of mutation breeding where plants
are exposed to radioactive sources, in order to generate
useful mutations.

33.  Possible to achieve instant progress in elite material.
 Single trait improvements can be made to an established variety
preferred by producers, processors and/or consumers.
 Limited breeding effort required and treated material is safe to handle.
 Novel variation can be produced.
 Single gene mutants with no negative pleiotropic effects are possible.
 For some mutagenic treatments such as gamma and X-ray, there is
neither residual radiation nor chemical contamination of the treated
material.
 Specific genes/traits can be targeted.
 Possible to calculate chances of success (mutation frequency).
Advantages

34.  The frequency of desirable mutants is very low.
 The process is generally random and unpredictable.
 Identification of micro mutation, which are more useful to a plant
breeder is usually very difficult.
 Mutants have strong negative pleiotropic effects on other traits.
 Health risks: handling, chemical mutagens; radiations, fast neutrons
treatments.
Ways to Mitigate These Limitations-
 We can mitigate these problems of unpredictable fate of mutations
by increasing the size of population so as to get a good number of
mutants.
 Efficient handling of the mutagens should we focused on to eradicate
any health hazards if any.
Disadvantages

35. More than 3,200 mutant varieties including numerous crops
(75 %), ornamentals and trees (25 %) have officially been
released for commercial use in more than 210 plant species
from over 70 countries.
(Source: fao/ iaea mutant varieties database) .
PERCENTAGE OF MUTANT VARIETIES BY CROP TYPE
Contd…

37. Data source: FAO/IAEA Mutant variety database, 2018

38. S. No Mutagen used Number of released
mutant cultivars
1. Gamma rays 910
2. X-rays 311
3. Gamma chronic 61
4. Fast neutrons 48
5. Thermal neutrons 22
6. Ethyl methyl sulphonate 106
7. Sodium azide 11
8. N-ethyl-N-Nitroso urea 57
9. N-Nitroso-N-methyl urea 46
Data source: FAO, 2015
Number of officially released mutant cultivars with different
types of mutagens

39.  Higher yield Barley (DL 253), Pea (Hans)
 Groundnut (Co 2, TG 17)
 Short stature Barley (RDB 1),Rice (Prabhavati)
 Earliness Rice (IIT 48,IIT 60,Indira,Padmini)
 Stress resistance Salt tolerance in Rice (Mohan)
 Water logging tolerance in jute ( Padma)
 Bold seed size Groundnut (PB 1,PB 2,Vikram) and
 Rice ( Jagannath)
Achievements of mutation breeding

40. Crop species Mutant var. Release year Mutagen
Cotton M.A.-9 1948 Gamma rays
Wheat NP-836 1961 X-rays
Sugarcane Co-6608 1966 Gamma rays
Castor Aruna 1969 Neutrons
Mulberry S-54 1974 EMS
Bougainvillea Arjuna 1976 Gamma rays
Blackgram Co-4 1978 MMS
Hibiscus Purnima 1979 Gamma rays
Sesame Kalika 1980 EMS
White jute Shyamali 1980 X-rays
Lentil S-256 1981 Radiation
Cowpea V-16 Amba 1981 DMS
Mungbean TAP-7 1981 Gamma rays
Chickpea WCG 2 1999 Gamma rays
World First Mutant Cultivars Released In India

41.  Improving specific characteristics of an otherwise
well adapted high yielding variety
 Used to improve various quantitative characters,
including yield.
 F1 hybrids from intervarietal crosses may be treated
with mutagens in order to increase genetic variability
by inducing mutations and by facilitating
recombinations among linked genes.
Application of mutation breeding in crop improvement

42. CASE
STUDIES

43. Inducing Potential Mutants in Bread Wheat Using Different
Doses of Certain Physical and Chemical Mutagens.
Case study -1
(Abaza et al., 2020)

44. 44
Case study -2
Improvement of Grain Yield and Yield Associated Traits in Bread Wheat
(Triticum aestivum L.) Genotypes Through Mutation Breeding Using
Gamma Irradiation.
(Balken et al .,2020)

45. Mutation breeding has become increasingly popular in
recent times as an effective tool for crop improvement.
The direct use of mutation in the development
of molecular maps in structural and functional
genomics could lead to rapid improvement of
plant yield and quality.
Mutation in association with the new
technology of genetic engineering will
constitute tools of plant breeders in near
future.
WAY FORWARD
1
2
3

46.  At present genetic variability is narrowed using conventional
breeding approaches for a long period, induced mutagenesis are
one of the most important approaches for broadening the genetic
variation and diversity in crops to circumvent the bottleneck
conditions.
 Induced mutagenesis, albeit can contribute to unleashing the
potentials of plant genetic resources and thereby avail plant
breeders the raw materials required to generate the envisaged
smart crop varieties.
 Crop varieties generated through the exploitations of mutation
breeding are significantly contributing to global food and
nutritional security and improved livelihoods.
CONCLUSION