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利用根癌土壤杆菌 (Agrobacteriumtumefaciens (SmithetTownsend)Conn)介导法将广谱抗虫基因豇豆胰蛋白酶抑制剂 (CpTI)基因导入毛白杨 (PopulustomentosaCarr.) 12 85雌株和毛新杨 (P .tomentosa×P .bolleana)×毛白杨回交杂种 ,比较了不同卡那霉素 (Km)浓度和Km选择压力的施加方式对毛白杨芽的再生过程和转化频率的影响 ,经过严格筛选获得了大量转基因株系。对所获得的转基因株系从试管苗继代能力、叶片再生芽能力和嫩茎生根能力 3个指标进行了卡那霉素抗性 (Kmr)鉴定 ,将Kmr 植株初步确定为转基因植株。对部分Kmr 植株进行了分子检测 ,PCR鉴定和PCR_Southern检测证实 ,CpTI基因已整合进杨树基因组中。CpTI抑制活性检测表明 ,大部分转基因株系都对胰蛋白酶有一定的抑制活性 ,对照未转化植株没有抑制活性。
A broad-spectrum insect-resistant gene cowpea trypsin inhibitor (CpTI) gene was introduced into 1285 female Populus treponementosa Carr. And P. somentosa × using the Agrobacterium tumefaciens (Smithetownown) Conn mediated method. P.bolleana × backcross hybrids of Populus tomentosa to compare the different kanamycin (Km) concentration and Km selection pressure on the regeneration process and transformation frequency of Populus tomentosa, after a rigorous screening of a large number of transgenic lines system. The transgenic lines were identified as kanamycin resistance (Kmr) using three indicators of subculture ability, regeneration ability of buds and rooting ability of young stems. Kmr plants were initially identified as transgenic plants. Some Kmr plants were subjected to molecular detection. PCR and PCR Southern detection confirmed that the CpTI gene was integrated into the poplar genome. CpTI inhibitory activity test showed that most of the transgenic lines have a certain inhibitory activity of trypsin, the control untransformed plants did not inhibit the activity.