Molecular farming involves genetically modifying crops to produce proteins and chemicals for medicinal and commercial use. It has several advantages over traditional pharmaceutical production methods, including lower costs and the ability to produce industrial-scale amounts of proteins. Molecular farming systems include stable nuclear and plastid transformation of crops, as well as transient transformation and hydroponic growth of transgenic plants. Edible vaccines produced in crops through molecular farming could provide mucosal immunity at low cost but require careful monitoring to ensure biosafety.

1. Molecular farming:
The Next destination for agriculture
biotechnology
MUTTURAJ M. D.
M.Sc.(GPB)
MA1TAF0179
3/24/2018 Dept. of GPB 1

2. Introduction
Costly medicines
3/24/2018 Dept. of GPB 2

3. Molecular farming
 Molecular farming is an experimental application of
biotechnology that involves the genetic modification
of crops for the production of proteins and chemicals
for medicinal and commercial purposes.
3/24/2018 Dept. of GPB 3Ahmad,2014

4. Brief history of molecular farming
Schillberg et al.,20133/24/2018 Dept. of GPB 4

5. Molecular Farming Hosts
 Bacteria
 Yeasts, (single celled fungi)
 Unicellular algae
 Whole plants, ( corn, barley, rice, duckweed, tobacco)
 Whole animals, (insects, birds, fish, mammals)
3/24/2018 Dept. of GPB 5

6. Comparison of expression system
Ahmad,2014
3/24/2018 Dept. of GPB 6

7. Advantages Plant Molecular Farming
Daniell et al.,20013/24/2018 Dept. of GPB 7
Amount of recombinant product = industrial-scale levels.

8. Simplified Representation of Molecular Farming
3/24/2018 Dept. of GPB 8

9. Molecular farming systems
1.Stable nuclear transformation of a crop species
that will be grown in the field or a greenhouse
2.Stable plastid transformation of a crop species
3.Transient transformation of a crop species
4.Stable transformation of a plant species that is
grown hydroponically
Horn et al.,20043/24/2018 Dept. of GPB 9

10. 1.Stable nuclear transformation of a crop species that
will be grown in the field or a greenhouse
3/24/2018 Dept. of GPB 10

11. Advantages
 Long term non refrigerated storage.
 Large acreage can be utilized with the lowest possible cost.
Disadvantages
 Outcrossing
 Higher manual labour requirements
 Lower yields
Horn et al.,20043/24/2018 Dept. of GPB 11

12. 2.Stable plastid transformation of a crop species
3/24/2018 Dept. of GPB 12

13. Advantages
 Higher yield of foreign protein than nuclear transformation.
 Plastid genes are not usually transmitted through pollen so
that outcrossing is not a major concern.
Disadvantages
 Protein stability over time will change even with refrigeration.
 Extraction and purification must be performed at very specific
times .
 Large volume products and edible vaccines would not appear
to be feasible using this system.
Horn et al.,20043/24/2018 Dept. of GPB 13

14.  The WHO recommends complete withdrawal of oral polio
vaccine (OPV) by April 2016 globally and replacing with
at least one dose of inactivated poliovirus vaccine (IPV).
Chan et al.,20163/24/2018 Dept. of GPB 14

15. 1. VP1 chloroplast vector construction and regeneration of
transplastomic plants
Integration of VP1 into tobacco
and lettuce plastomes
 To confirm transgene integration into
the tobacco chloroplast genome, PCR
was performed using primer.
 Southern blot analysis was performed to
confirm transgene integration.
Chan et al.,2016
3/24/2018 Dept. of GPB 15

16. • Lyophilization of CTB-VP1
transplastomic tobacco leaves.
• Purification of chloroplast
derived proteins.
Immunization schedule
G1: control
G2-G8: first IPV
G2-G3: Boosted with second IPV
G4-G10: boosted with
lyophilized plant material.
The strategy of using a plant-made VP1
subunit vaccine for an oral booster rather
than repeated OPV vaccination is a novel
approach to achieve the goal of global
poliovirus (PV) eradication.
Chan et al.,20163/24/2018 Dept. of GPB 16

17. 3.Transient transformation of a crop species
3/24/2018 Dept. of GPB 17

18. Advantages
 Small quantities of the target protein can be obtained within
several weeks.
 Probably the ideal system for a large number of custom
proteins that are needed in relatively small amounts.
Disadvantages
 Probably not suitable for any protein needed in large (kg)
quantities.
 Product must be processed immediately as storage will cause
degradation of the plant tissue.
Horn et al.,20043/24/2018 Dept. of GPB 18

19. 4.Stable transformation of a plant species that is grown
hydroponically
 Transgenic plants containing a gene coding for the target
protein are grown hydroponically in a way that allows release
of the desired product as part of the root exudate into the
hydroponic medium3/24/2018 Dept. of GPB 19

20. Advantages
 Plants grown hydroponically are contained in a greenhouse
setting and so have reduced fears of unintentional
environmental release.
 Purification of the desired product is considerably easier since
no tissue disruption is needed and the quantity of
contaminating proteins is low.
Disadvantages
 Probably not amenable to producing large (kg) quantities of
any protein product.
 Greenhouse/hydroponic facilities are relatively expensive to
operate.
Horn et al.,20043/24/2018 Dept. of GPB 20

21. Proteins of Molecular Farming
Parental therapeutics
and pharmaceutical
intermediates
Industrial proteins
and enzymes
Monoclonal
antibodies
 This group includes all proteins used directly as pharmaceuticals along with
those proteins used in the making of pharmaceuticals.
 Includes such products as thrombin and collagen (therapeutics), and trypsin
and aprotinin (intermediates).
 This group includes hydrolases (glycosidase and proteases).
 Enzymes involved in biomass conversion for the purposes of
producing ethanol are candidates for molecular farming.
 This group includes all antibody forms (IgA, IgG, IgM,
secretory IgA, etc.).
 Plantibodies.
3/24/2018 Dept. of GPB 21

22. Edible vaccines
• A vaccine is a biological preparation that provides
active acquired immunity to a particular disease.
• A vaccine typically contains an agent that resembles a disease-
causing microorganism and is often made from weakened or
killed forms of the microbe.
Why we need vaccine ?
• Vaccines will help keep you healthy.
• Immunizations can save your child’s life.
• Vaccination is the most effective way to prevent infection and
severe outcomes caused by some viruses.
3/24/2018 Dept. of GPB 22

23. 3/24/2018 Dept. of GPB 23

24. Lettuce
Protein is extracted and analysis3/24/2018 Dept. of GPB 24

25. Immunization of human volunteers
• Five adult volunteers (male and female), 25–59 years
of age and in good health.
• The volunteers had no previous HBV vaccination, no
history of HBV infection, and no detectable anti-HBs.
• Three individuals received transgenic lettuce leaves
twice: 200 g at first, and within 2 month, 150 g.
• Two control individuals were given the same amount
of non-transgenic lettuce.
Kapusta et al.,19993/24/2018 Dept. of GPB 25
Humans may be immunized orally against
HBV with plant expressing viral antigen

26. Advantages of Edible Vaccines
• An effective means for immunization .
• Elicit mucosal immunity unlike the conventional vaccines.
• Economical in terms of cost of production , storage and delivery.
• They find easy acceptance.
• Provide immunization against several diseases.
Shah et al., 2011
3/24/2018 Dept. of GPB 26

27. Examples of Edible Vaccine
Neeraj et al.,2008
3/24/2018 Dept. of GPB 27

28. Bio-safety
• Contamination of the food chain due to outcrossing.
• Using the same equipment for harvesting and
processing of transgenic and food crops.
• Growing food crops in the same field where a
transgenic crop was grown previously.
3/24/2018 Dept. of GPB 28

29. Monitoring
• To avoid the food chain contamination, Geographically
isolating the transgenic crop, growing in greenhouses
instead of open field.
• Harvesting and processing transgenic plants using separate
equipment or properly decontaminating the equipment.
• Use of male sterility system, the chances of transgene
spread through pollen are controlled.
• It is also important to label genetically modified products so
that the consumer has the choice to select according to his/
her own preferences.
3/24/2018 Dept. of GPB 29

30. 3/24/2018 Dept. of GPB 30