This presentation highlights the uses of transgenic plants as bio-pharamaceuticals including plantibodies, edible vaccine etc. The mechanism of plantibodies, vaccine generation has also been focused

1.  Cheaper than mammalian, insect, or
prokaryotic systems
 Agricultural infrastructure already in place
 Easy to scale-up
 No human pathogens, endotoxins in plants
 Correct folding, modifications
 High volume
 Direct use
Dr. Soumitra Paul, MPP lab, Dept. of
Botany, C.U.

2. Humanitarian:
Improved
farming
Cheaper
food
More food
Increased nutrition
Reducing
world
hunger
and
improving
world
health
Pest resistance
Herbicide
resistance
Cold tolerance
Drought
tolerance
Edible vaccines
Environmental: reduced use of herbicides
and chemicals in farming. Dr. Soumitra Paul, MPP lab, Dept. of
Botany, C.U.

3.  Tissue-specific expression
- Oil bodies, seeds, tubers
 Plastid transformation
 Molecular approaches
 Protein targeting
 Higher yield
 Long-term stability
 Reduced toxicity
 Correct processing
Dr. Soumitra Paul, MPP lab, Dept. of
Botany, C.U.

4. Dr. Soumitra Paul, MPP lab, Dept. of
Botany, C.U.

5.  Targeting information is contained in peptide
signaling sequences
 Signals are recognized by receptors that direct
them to the target organelle
 Translocation via:
 Transmembrane channels
 Vesicles
Dr. Soumitra Paul, MPP lab, Dept. of
Botany, C.U.

6. Protein Targeting: examples & mechanisms:
Via cloning we create fusion proteins that direct your
protein to the desired location.
The signal peptides come from previously identified
genes.
The primary locations for targeting are members of the
endomembrane system (ER, Golgi, Vacuole) and outside
of the cell (in plants the apoplast).
Experimentally how do we do this??
Dr. Soumitra Paul, MPP lab, Dept. of
Botany, C.U.

7. Arabidopsis protoplast
Red = chloroplasts;
Green = mitochondria
Mitochondrian Chloroplast
For example organelles
Arabidopsis leaf
vacuole
Dr. Soumitra Paul, MPP lab, Dept. of
Botany, C.U.

8.  Apoplast
Default pathway
 Endoplasmic reticulum
Conserved C-terminal H/KDEL sequence
Retention/recovery signal
ER TS
-HDELER TS
Dr. Soumitra Paul, MPP lab, Dept. of
Botany, C.U.

9.  The endoplasmic reticulum target signal
 N-terminal
 Conserved structure
 Directs co-translational import into the
endoplasmic reticulum
Dr.SoumitraPaul,MPPlab,Dept.of
Botany,C.U.

10.  Vacuole
 Sequence Specific Vacuolar Sorting Determinant
Typically N-terminal
Conserved NPIR consensus sequence
 C-Terminal Vacuolar Sorting Determinant
No conserved sequence
Hydrophobicity important
ctVSDER TS
ssVSDER TS
Dr. Soumitra Paul, MPP lab, Dept. of
Botany, C.U.

11.  N-terminal
transit signal
 Imported into
stroma post-
translationally
 May have
additional
signals for
further sorting
Jarvis and Robinson 2004Dr. Soumitra Paul, MPP lab, Dept. of
Botany, C.U.

12.  Benefits:
 Higher yields
 Post-translationally
modified
 Example:
 Accumulation of phytase in tobacco leaves
(Verwoerd et al. 1995)
Reached levels of 14.4% phytase per mg of total
soluble protein
Cry operon of three genes showed insecticide
resistance in transplastomic tobacco
Dr. Soumitra Paul, MPP lab, Dept. of
Botany, C.U.

13. Plantibodies:
Application of Plant molecular breeding:
molecular pharming
Transgenic plants produce functional monoclonal antibodies,
inexpensive methods.
Target region: Seeds (Cereals, legumes), Tubers (Potato)
Type:
Single chain functional variable protein (scFv). Eg. scFv of
IgG of mouse B-cells in tobacco for Hodgkins lymphoma
Secretary immunoglobulin A (sIgA) for Streptococcus mutans
in tobacco (dental caries)
Full length IgG1 for S mutans. Dr. Soumitra Paul, MPP lab, Dept. of
Botany, C.U.

14. Vaccine developed in transgenic plants..
Factors:
i. Stimulate mucosal membrane and first target for oral vaccine is the
diarrheal diseases
ii. Ordered structures resistant to digestion
iii. As foreign antigen (epitope) for production of antibodies
iv. Should be edible part..
Tobacco is not used
later Why?????1. Hepatitis B in tobacco/Carrot/banana
2. Autoimmune diabetes in potato
3. HIV in tobacco/bean
4. Rhinovirus in bean
5. Cholera and E. coli diarrhoea in potato
Two ways:
1. Structural gene into plant transformation vector,
2. Chimeric with plant viral coat protein gene
Edible Vaccine:
Dr.SoumitraPaul,MPPlab,Dept.of
Botany,C.U.

15. Biopharmaceuticals:
1. Anticoagulants: Hirudin in oilseed rape
2. Recombinant hormones: Human interferon in rice
3. Protein/ peptide inhibitors: Human α 1 antitrypsin in rice
4. Recombinanat enzymes: Glucocerebrosidase in tobacco,
Gaucher’s disease
5. Human serum albumin in tobacco
6. Cyclodextrins (CGTase from Klebsiella) in potato
Dr. Soumitra Paul, MPP lab, Dept. of
Botany, C.U.