Efficient production of human acidic fibroblast growth factor in pea (Pisum sativum L.) plants by agroinfection of germinated seeds

Background For efficient and large scale production of recombinant proteins in plants transient expression by agroinfection has a number of advantages over stable transformation. Simple manipulationپCrapid analysis and high expression efficiency are possible. In pea, Pisum sativum, a Virus Induced Gene Silencing System using the pea early browning virus has been converted into an efficient agroinfection system by converting the two RNA genomes of the virus into binary expression vectors for Agrobacterium transformation. Results By vacuum infiltration (0.08 Mpa, 1 min) of germinating pea seeds with 2-3 cm roots with Agrobacteria carrying the binary vectors, expression of the gene for Green Fluorescent Protein as marker and the gene for the human acidic fibroblast growth factor (aFGF) was obtained in 80% of the infiltrated developing seedlings. Maximal production of the recombinant proteins was achieved 12-15 days after infiltration. Conclusions Compared to the leaf injection method vacuum infiltration of germinated seeds is highly efficient allowing large scale production of plants transiently expressing recombinant proteins. The production cycle of plants for harvesting the recombinant protein was shortened from 30 days for leaf injection to 15 days by applying vacuum infiltration. The synthesized aFGF was purified by heparin-affinity chromatography and its mitogenic activity on NIH 3T3 cells confirmed to be similar to a commercial product. Background The acidic mammalian fibroblast growth factor (aFGF) and the basic fibroblast growth factor (bFGF) bind to heparin decasaccharide and to domains of their tyrosine membrane spanning kinase receptor [1, 2]. FGF is a powerful mitogen in many mammalian cell types. However its major importance is to switch endothelial cell growth to angiogenesis (formation of blood vessels) and to development of tumors [3, 4]. Depending on the cell growth substrate FGF either stimulates endothelial cell growth or promotes capillary differentiation. Extensive cell spreading and growth were stimulated when the culture dishes were pre-coated