Lecture 2. Transgenic Plants Viral Vectors Transient Expression. Expression in Transgenic Plants. Limitations some crops can not be regenerated vulnerable to weather not 100% biologically contained unless via chloroplast transformation. Advantages low initial investment
Viral Vectors for the expression of proteins in plants
Yuri Gleba, V. Klimyuk and S. Mariollnnet
Current Opinioins in Biotech 2007 18: 134-141
Large Scale Biology Corp.
Vacuum – 0.8 bar for 20 sec
Protein A fragments
Gene encoding the desired protein.
Incorporate into a plant transformation vector for optimized expression in plant cells.
Stable expression: Nuclear genome integration.
Stable expression: Chloroplast genome integration.
Integrate into a viral coding sequence for expression as a “by product” of viral replication.
Transient expression: Infect plant to initiate viral replication.
Modify viral genome to adapt it into a plant transformation vector for subsequent regulated release as a replicon in transgenic plants.
Primarily for antigens/epitopes until very recently
Identify protective epitope within antigenic protein.
Create viral replicon coding sequence with epitope fusions to coat protein.
gene of interest
pA35SpTRA Plant Expression Vector
Agrobacterium-infiltration as a useful tool for rapid assessment of gene expression in plants
Gene of Interest
wt 945(P) A 945 (A)
1 2 2d 5d
Plant Vaccine Group University of Cape Town
Optimising protein production in plants
1. Select optimal constructs using agroinfiltration - simple and rapid method for evaluating protein expression of numerous Agrobacterium constructs
2. Small OR Large-scale infiltration