TISSUE CULTURE OF AMERICAN GINGSENG AND GENETIC ENGINEERING TO EXPRESS ANTIFUNGAL PROTEINS

Transformation of American ginseng (Panax quinquefolius L.) with Agrobacterium strain LBA 4404 containing either a rice chitinase gene or a thaumatin-like protein gene under control of the maize ubiquitin1 promoter was achieved.
The phosphinothricin acetyltransferase (bar) and hygromycin phosphotransferase (hpt) genes were used as selectable markers.
Epicotyl explants from 2 to 3-week-old ginseng seedlings were pre-cultured for 5-7 days on MS medium with NAA and 2,4-D at 10 μM and 9 μM, respectively (ND medium), prior to Agrobacterium infection.
The explants were either immersed in a bacterial suspension for 20 min or received a 10-15 μL droplet of bacteria.
Following this, a co-culture period of 3-4 days was provided on ND medium.
Selection was conducted using 20 mg L^-1 phosphinothricin or 100 mg L^-1 hygromycin over a 10-month period.
A callusing frequency of 24-27% was achieved on ND medium when explants were infected by the droplet method.
Immersion of explants reduced the callusing frequency to 9.3%. Almost 90% of 32 lines that survived selection were shown to be transformed by polymerase chain reaction (PCR) analysis.
The expression of the chitinase and TLP genes was demonstrated by Western analysis.
One hundred and two ginseng plantlets were recovered from somatic embryos of 11 confirmed transgenic lines.
The transgene integration in plantlets of two lines was also demonstrated.
This report of Agrobacterium-mediated transformation of an important medicinal plant has the potential to enhance tolerance to fungal diseases.