유전자총 실험조건 최적화를 통한 형질전환 백합 식물체 생산

Optimization of a protocol for the production of transgenic lily plants via particle bombardment

Transgenic lily plants have been obtained after particle bombardment, using PDS-1000/He system and scale explants of lilies, followed by PPT (D-L-phosphinothricin) selection. In this study, scales of the lily plants cv. `red flame` were bombarded with a plasmid containing the bar gene as a selectable marker, and the AtSIZ gene as a gene of interest, showing salt tolerance and drought tolerance respectively, and both being driven by the CaMV 35S promoter. For optimization of a protocol, factors which optimized and showed a high transformation efficiency under following conditions, were considered: a bombardment pressure of 1100 psi, a target distance of 6 cm and 1.0 μm of gold particle, and 24-h pre-culture and post-culture on MS medium containing 0.2 M sorbitol and 0.2 M mannitol as osmoticum agents. After bombardment, all the bombarded scales of lily were transferred to MS medium without selective agents, for a week. Subsequently, these bombarded scales were transferred to a selection MS medium containing 10 mg/l PPT, and incubated for a month for further selection, after which they were cultured for another 4-8 weeks with a 4-week subculture regime on the same selection medium. After transferring into hormone-free MS medium, the PPT-resistant scales with shoots were successfully rooted and regenerated into plantlets. PCR analysis revealed that the surviving putatively transformed plantlets indicated the presence of both the bar gene and the AtSIZ gene. In conclusion, when 100 scales of lil cv. Red flame are bombarded, this study produced approximately 17-18 transgenic plantlets with an optimized bombardment protocol. The protocol described here can contribute to the breeding program of lilies.