Agrobacterium-mediated transient expression may be regarded as a promising method for inexpensive large-scale production of recombinant proteins. We optimized the protocol of transient expression in Nicotiana benthamiana and compared six Australian species of Nicotiana as hosts for transient expression. The transient expression of GFP under 35S CaMV promoter was observed in all species tested, although the GFP content in leaves of N. benthamiana, N. exigua, and N. excelsior was significantly higher (3.8, 3.7, and 2.0% TSP, respectively). Usage of viral-based expression system resulted in considerable increase of GFP accumulation in N. excelsior and N. benthamiana (63.5 and 16.2% TSP, respectively). We displayed that N. excelsior has the best characteristics in regard to biomass yield as well as GFP accumulation level for both types of the expression cassettes tested.
In vitro production of the meroterpene bakuchiol by Psoralea drupacea Bge (Fabaceae) has been studied using aseptically-grown plants, callus cultures of different origin, cell suspensions and transgenic hairy root cultures. The effect of phytohormones and methyl jasmonate on bakuchiol production was also investigated. Bakuchiol was not detected in cell suspensions or hairy root preparations of P. drupacea. In contrast, aerial parts of P. drupacea grown in vitro were found to accumulate up to 11% dry weight of bakuchiol and can therefore be regarded as a potentially useful source of this antimicrobial compound.
In this study, we report the obtaining of carrot plants expressing human interferon alpha-2b via Agrobacterium-mediated transformation using two vector constructs containing the sequence coding for interferon gene fused with Nicotiana plumbagenifolia calreticulin apoplast targeting signal driven by 35S CaMV promoter and root-specific Mll promoter. The human interferon alpha-2b gene was correctly translated in carrot plants according to Western blot analysis. The recombinant protein exhibited antiviral activity in vitro by inhibition of vesicular stomatitis virus replication in established piglet testicular cells. The results demonstrated the higher activity of interferon accumulated in carrot plants for young leaves (up to 50.7 × 10(3) IU/g FW) compared to the mature ones probably due to the degradation-susceptible nature of this protein. The taproot-expressing system could have also provided the sufficient protein amounts (up to 16.5 × 10(3) IU/g FW) and could possibly be used for generating interferon alpha-2b protein in planta for preventing and curing infectious diseases.
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