Producing disease free plants in vitro means

1. Producing disease free
plants using in vitro means
Presented by: Sarah Ashfaq
Roll No. : 51
BS 7TH Semester ADP Replica
Institute of Botany, University of the Punjab
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2. Contents
1. Introduction
2. In vitro technique
3. Production of disease free plants
4. Virus eradication methods
5. Role of Green house
6. Conclusion
7. References
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3. Introduction
• Plant tissue culture is a method or technique to isolate parts of
plants (Protoplasm,cells,tissues and organs) and grow them on an
artificial media so that parts of these plants can grow and develop
into complete plants.
• This culture is on nutrient media under sterile conditions.Pierik
et al., (1997).
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4. In Vitro Technique
• Culture in vitro is used to obtain disease-free plants and rapid
multiplication of the new clones.
• In vitro techniques were established around 1980.
• Italian, French and Spanish commercial firms are
selling artichoke plants propagated in vitro.
• These cultures are initiated from shoot apex.
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5. Production of disease free plants
• Plants infected by bacteria and fungi may respond to treatments
with bactericidal and fungicidal compounds there is no
commercially available treatment to cure virus infected plants.
• Virus free plants can Produce by plant tissue culture.
• Selection can made it possible to choose healthy plant material if
not all crop is infected with virus.
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6. Virus Eradication Methods
Thermotherapy
Meristem tip
culture
Chemotherapy
In vitro shoot
tip grafting
Callus Culture
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7. Thermotherapy
High Temperature
• Parent plants materials are grown in
a specially designed growth
chamber at increased temperatures
of 30-40 Celsius for 2-3 months.
Low Temperature
• Low Temp 10-15 Celsius followed
by meristem culture has been
employed for production of virus
free plants.
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8. 7/5/2022 8
Host Virus eliminated Temperature
Chrysanthemum Chrysanthemum B virus 35-38◦ C
Carnation Carnation ringspot virus 35-40◦ C
Banana Cucumber mosaic virus 35-43◦ C
Goose berry Goose berry vein binding
virus
35◦ C
Potato Potato virus S,Y,Z 33-38◦ C
Examples

9. Meristem Tip Culture
• Explant:
• Shoot tip: Apex+ Leaf primordia ( 100-500 µm)
• Tip Meristem : (100 µm in diameter and 250 in length)
• Technique:
1. Too small size to be prepared.
2. With the help of microscope.
3. Prevent desiccation.
4. Care of plant before the collection of explant.
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10. Factors
Culture
medium.
Explant size.
Storage
condition.
Physiological
condition of
explant.
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11. Meristem tip culture
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Source:
https://biologyreader.com/meris
tem-culture.html

12. Chemotherapy
“The use of chemicals
to suppress virus
symptoms and
multiplication in plants.”
Use of antiviral
compounds.
Growth promoting
chemicals.
Antimetabolite
chemicals.
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13. 7/5/2022 13
Fig.Growth of in vitro cassava plantlets on Murashige and Skoog (MS) medium
after 42 days on ribavirin treatment. Control 0 mg/L (a), 10 mg/L (b), 20 mg/L (c)

14. Virus elimination through in vitro shoot tip
grafting
• Sometimes the shoots developed in shoot tip cultures do not forms roots.
• In some such cases it may be possible to graft the shoots onto a virus free
rootstock derived from seedlings.
• This technique first used for Dahlia.
• Murashige et al., (1972) were able to obtain a few citrus plants by grafting
shoot tips from diseased plants on young rootstock seedlings growing in
vitro.
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15. 7/5/2022 15
Fig. Microcuttings of dahlia
obtained from shoot tip explants
after 8 weeks of cultivation on MS
media
Source:
Source:https://www.researchgate.n
et/figure/Microcuttings-of-dahlia-
obtained-from-shoot-tip-explants-
after-8-weeks-of-cultivation-
on_fig1_337248017

16. Virus elimination through callus culture
• “Production and maintenance of callus on a solidified nutrient medium under
aseptic condition”.
Steps of callus culture
1. Selection and collection of explant
2. Surface sterilization
3. Preparation of explant
4. Inocubation of culture
5. Subculture of callus
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17. Cont…
• Regeneration of plant from callus
1. Somatic embryogenesis
2. Organogenesis
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18. 7/5/2022 18
Fig. Callus culture various
steps
Source: Sarah Ashfaq

19. Conclusion
• From the various example given in this topic it is apparent that tissue culture
techniques have a useful role in the eradication of systematic disease in
plants caused by viruses.
• Meristem tip culture seems the most reliable method for pathogen
elimination.
• In vitro shows a better rooting ability after planting out.
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20. References
• Introduction to Plant Biotechnology By H. S. Chawla.
• Perspectives in Plant Cell and Tissue Culture By I. K. Vasil.
• Plant Tissue Culture: Theory and Practice By S.S. Bhojwani, M.K.
Razdan.
• Wang, P. J., and C. Y. Hu. "Regeneration of virus-free plants through in
vitro culture." Advances in Biomedical Engineering, Volume 18. Springer,
Berlin, Heidelberg, 1980. 61-99.
• Pierik, Rudolf Leonardus Maria. In vitro culture of higher plants. Springer
science & business media, 1997.
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Thankyou