1. Direct embryogenesis In direct somatic embryogenesis, the embryo is formed directly from a cell or small group of cells without the production of an intervening callus. 2. Indirect embryogenesis In indirect somatic embryogenesis, callus is first produced from the explant. Embryos can then be produced from the callus tissue or from a cell suspension produced from that callus.

1. PBG – 604 – Molecular and Chromosomal Manipulation in Crop Improvement
(2+0)
Topic - Organogenesis and somatic embryogenesis
In vitro mutant selection for biotic and abiotic stresses
Department of Plant Breeding and
Genetics
Presented By -
Jyoti Prakash Sahoo
01ABT/PHD/17
Dept. of Agril. Biotech.
OUAT, BBSR
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2. Organogenesi
s
 The formation of roots, shoots or flower buds from the cells in culture
in manner similar to adventitious root or shoot formation in cuttings is
called organogenesis.
1. Direct regeneration
-
2. Indirect organogenesis -
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3. Somatic
Embryogenesis
1. Direct embryogenesis
 In direct somatic embryogenesis, the embryo is formed directly from a cell or
small group of cells without the production of an intervening callus.
2. Indirect embryogenesis
 In indirect somatic embryogenesis, callus is first produced from the explant.
 Embryos can then be produced from the callus tissue or from a cell
suspension produced from that callus.
 The process of a single cell or a group of cells initiating the developmental
pathway that leads to reproducible regeneration of non-zygotic embryos
capable of germinating to form complete plants.
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4. PhysicalAgents
Typeof Mutation
Induction
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Ionizing radiations - such as X-rays, gamma rays and alpha particles cause
DNA breakage. The most common lab sources include cobalt-60 and cesium-137.
Ultraviolet radiations with wavelength above 260 nm are absorbed strongly by
bases, producing pyrimidine dimers, which can cause error in replication if left
uncorrected.
Radioactive decay, such as 14C in DNA which decays into nitrogen.

5. 1.Baseanalogs - bromouracil(BU),aminopurine (AP)
2.Chemicalswhichalter structureandpairing propertiesofbases
Alkylating agents [ethyl methanesulphonate (EMS); diethyl
sulphate (dES); ethyleneimine (EI); ethyl nitroso urethane
(ENU), ethyl nitroso urea (ENH), methyl nitroso urea
(MNH)
3.Intercalatingagents
Acridineorange,Proflavin,Ethidium Bromide
ChemicalMutagens
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6. 6
BiologicalAgents
Transposon, a section of DNA that undergoes autonomous fragment
relocation/multiplication. Its insertion into chromosomal DNA disrupts
functional elements of the genes.
Virus – Virus DNA may be inserted into the genome and disrupts
genetic function.
Bacteria – Some bacteria such as Helicobacter pylori cause
inflammation during which oxidative species are produced, causing DNA
damage and reducing efficiency of DNA repair systems, thereby
increasing mutation.

7. Site Directed Mutagenesis
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8. 8
Oligonucleotide-directed mutagenesis

9. 9
Mechanism of somaclonal variation in
micropropagated plants as a result of oxidative
burst upon in vitro culture

10. Invitro Mutant Selection For Salt
Tolerance
 Generally an array of stresses interplay in saline soils and reduces productivity
of salt sensitive crops.
 The optimum salt concentration of the growth of halophytes is found to be
about 0.5 M Nacl .
Injury Mechanism
• Reduce water potential
• Ionic imbalance
• Inhibition of enzymatic imbalance
• Change in physiological
parameter
• Inhibition of translocation
Possible Tolerance Mechanism
• Antioxidative defense system
• Ion Homeostasis
• Accumulation of compatible
solutes
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11. 11
Antioxidative defense system – Induced by Salinity/other
abiotic stress
 ROS – Reactive Oxygen Species
 Contineously genetated in Mitochondira, Peroxisome and
Cytoplasm
 Destroys the normal metabolism through oxidative damage
Composed of - superoxide radicals (O2−)
hydrogen peroxide (H2O2),
hydroxyl radicals (OH• )
singlet oxygen (1O2).
Plant cells have developed complex antioxidant defense system
 enzymatic (SOD, APX, GPX, GR, CAT, etc.)
 non-enzymatic (ascorbate, glutathione, -tocopherol, carotenoids, flavonoids,
etc.) to protect themselves against saltstress

12. 12
Antioxidative defense system – Induced by Salinity/other
abiotic stress
Pathways for ROS-scavenging in plant cell
(Mittler, 2002; Apel and Hirt, 2004)
Superoxide dismutase (SOD)
Ascorbate peroxidase (APX)
Catalases (CAT)
Glutathione reductase (GR
GSH(reduced
glutathione)
GSSG (oxidized
glutathione)
chloroplasts
chloroplast,
cytosol,
mitochondria,
apoplast and
peroxisomes
peroxisomes

13. 13
Ion
Homeostasis
S. Mahajan, N. Tuteja / Archives of Biochemistry and Biophysics 444 (2005)

14. 14
Mechanism of Stress
Tolerance
S. Mahajan, N. Tuteja / Archives of Biochemistry and Biophysics 444 (2005)

15. Screening of Salt
Tolerance
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16. 16
Screening and invito selection for abiotic stresses ( Drought, Low
Temp. and UV)

17. 17
Role of ROS under conditions of pathogen
attack (Biotic Stress)
Rapidly
dismutated
(Dangl and Jones, 2001; Apel and
receptor-induced signaling
reactive oxygen species

18. Inviro selection process for disease
resistance
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19. 19
Screening and invito selection for disease
resistance
Selecting agents – Culture Filtration (CF), DON – Deoxynivalenol, P- Pathogen, PT -
Pathotoxin

20. Plant Breeding Schemeby induced mutations and somaclonal
variation
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21. 21
References

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