Abstract
Calli were induced on MS medium supplemented with 0.5 mg/L 2,4-D by using the leaf explants of haploid which were derived from the diploid and haploid of Nicotiana tabacum cv BY4. These calli were subcultured on MS medium with the combination of 2.0 mg/L 2,4-D, 1.0 mg/L kinetin and 0.1 mg/L BAP. Cell propagation of diploid plants were good in a combination of 2.0 mg/L 2,4-D, 0.1mg/L BAP in vitro conditions, suspension cultures were conducted in equal condition. Homogenized suspension cultured cells were smeared 2.0 mL each on MS medium with 0~100 $\mu$M PFP, to select the resistant colony to PFP, and were examined after 10d, 20d and 30d. Measurment of fresh weight of cells after 30d of culture shows that with more concentration of PFP in medium the fresh weight of the cells decreased. In case of diploid, selected callus was the highest in vitro treated with 5 $\mu$M PFP. It was higher than control until 100 $\mu$M PFP. The active degree of catalase was the highest in vitro with 5 $\mu$M PFP but the lowest in vitro with 10 $\mu$M PFP on the other hand, in case of haploid plant, the active degree of peroxidase and catalase was the highest in vitro treated with 50 $\mu$M PFP. It's sure that enzyme active degree of between diploid and haploid had big differences.
담배 (Nicotiana tabacum cv. BY4)의 이배체 식물과 약배양으로부터 유도된 반수체 식물의 잎절편을 사용하여 0.5mg/L 2.4-D 단독처리구에서 캘러스를 유도한 후, 2.0 mg/L 2.4-D, 1.0 mg/L Kinetin 및 0.1 mg/L BAP를 조합처리하여 계대배양을 하였다. 이배체에서는 2.0 mg/L 2.4-D와 1.0 mg/L 카이네틴의 조합처리구에서, 반수체는 2.0 mg/L2.4-D 와 0.1 mg/L BAP 조합처리구에서 세포증식이 양호하였으므로, 현탁배양도 동일조건에서 실시하였다. 세포괴를 균일하게 한 현탁배양세포를 p-Fluorophenylalanine (PFP)이 5~100 $\mu$M의 농도로 단독처리한 배지상에 2.0 mL씩 평판배양하여, 10, 20, 30일 후 PFP의 처리에 대한 저항성 colony를 조사하였다. 배양 30일 후 세포주의 생중량을 측정한 결과 PFP의 농도가 높아질수록 감소하였다. 선발된 캘러스는 이배체서 peroxidase의 활성도가 5 $\mu$M PFP 처리시에 가장 높았는데 100 $\mu$M PFP 처리까지는 대조구보다는 높았다. Catalase의 활성도는 5 $\mu$M PFP 처리시 가장 높았으나 100 $\mu$M PFP 처리시 가장 낮았다. 한편 반수체에서는 peroxidase와 catalase의 활성도는 50 $\mu$M PFP 처리시 가장 높게 나타나서 이배체와 반수체 사이에 효소활성의 차이는 뚜렷했다.
