Abstract
Salt-tolerant transgenic Panax ginseng plants were produced by introducing the SAL1 geue (3'(2'), 5'-bis-phosphate nucleotidase) that confers tolerance to the salts through Agrobacterium tumefaciens co-cultivation. Cotyledon explants of immature ginseng zygotic embryos cultured on Murashige and Skoog medium lacking growth regulators formed somatic embryos directly with below 10%, but the 74% tranformation rate were observed at the treatment of phytohormone with 1.0 mg/l 2,4-D and 0.5 mg/l kinetin. Somatic embryos were initially cultured on MS medium supplemented with 250 mg/l cefotaxime for 3 weeks and subsequently subcultured five times to a medium containing 100 mg/l kanamycin and 250 mg/l cefotaxime. Upon development into the cotyledonary stage, these somatic embryos were transferred to on the medium containing 50 mg/l kanamycin and 10 mg/l gibberellic acid to induce germination and strong selection. Integration of the transgene into the plants was confirmed by polymerase chain reaction with specific primers. The ginseng transformants with well-developed shoots and roots were successfully acclimatized in a greenhouse when they were planted in soil.
인삼에 염류내성을 증진시키기 위해서 Arobidopsis에서 분리한 SAL1 (3‘(2’),5‘-bis-phosphate nucleotidase) 유전자를 Agrobacterium Tumefaciens을 이용하여 인삼자엽으로부터 형질 전환체를 유도하였다. Agrobacterium 과 공동배양 후 식물호르몬 무첨가 선발배지 (kanamycin 100 mg/l)에 치상한 결과 10%미만의 자엽에서 형질전환 인삼체세포배가 발생되었으나, Agrobacterium과 공동배양 후 1.0 mg/l 2.4-D와 0.5 mg/l kinetin의 식물호르몬을 첨가한 배지에 옮겨준 경우에는 74%의 형질전환율을 보였다. 발생한 체세포배는 초기에 250 mg/l 의 cefotaxime이 첨가된 MS배지에서 3주간 배양한 후 100 mg/l kanamycin과 250 mg/l cefotaxime이 첨가된 MS배지에 계대배양하여 선발하였다. 자엽단계로 발달한 체세포배들은 발아시키기 위해서 50 mg/l kanamycin과 10 mg/l 지베렐린이 첨가된 MS 배지로 옮겨 선발하였다. Kanamycin 첨가배지에서 선발된 체세포배들은 특이 프라이머로 PCR 증폭을 통하여 최종적으로 형질전환체를 확인하였으며, 줄기와 뿌리가 잘 발달된 형질전환체들은 성공적으로 토양에 순화시켰다.
