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通过PEG处理把外源基因导入甘蓝型油菜原生质体。转化介质中二价阳离子的种类和浓度、携带DNA及PEG溶液的pH值都会影响基因导入效率。以潮霉素抗性和卡那霉素抗性作标记,均成功地筛选到了抗性愈伤组织。相对转化频率分别为1.3%和0.2%。前者明显高于后者。把抗性愈伤组织转到分化培养基上,分化出芽。诱导生根后移栽到土壤中,生长状况良好。酶活性测定和Southern blotting分析表明外源基因已插入到植物细胞基因组中。该遗传转化系统存在的主要问题是抗性愈伤组织分化频率较低。本文对其原因作了初步分析。
Foreign genes were introduced into protoplasts of Brassica napus by PEG treatment. The type and concentration of divalent cations in the conversion medium, the pH of the DNA and PEG solution, will affect the efficiency of gene introduction. Resistance to hygromycin and kanamycin resistance were marked, were successfully screened to resistant callus. The relative transformation frequency was 1.3% and 0.2% respectively. The former is obviously higher than the latter. Resistant calluses were transferred to differentiation medium to differentiate buds. After induction rooting transplanted into the soil, the growth of good conditions. Enzyme activity assay and Southern blotting analysis showed that exogenous genes have been inserted into the plant cell genome. The major problem with this genetic transformation system is the low frequency of resistant callus differentiation. This article made a preliminary analysis of its causes.