摘要
以‘森田尼无核’葡萄为试材,研究了生长调节物质、外植体类型、抑菌素等对离体不定芽再生和转化效率的影响。相对于BA和KT,细胞分裂素TDZ无论单独使用还是与生长素配合使用均可使葡萄不定芽再生频率达到最高;住MS+TDZ 2.0 mg·L^(-1),MS+TDZ 4.0 mg·L^(-1)或MS+TDZ 4.0 mg·L^(-1)+IAA 0.1 mg·L^(-1)培养基上均可以较好地诱导再生不定芽;叶盘、茎段和叶柄都可以诱导再生不定芽,在同一种培养上再生频率差异不大;采用根癌农杆菌介导的叶盘法转化时,卡那霉素抗性筛选浓度为20 mg·L^(-1);抑菌抗生素使用头孢霉素适宜浓度为400 mg·L^(-1),而使用羧苄青霉素时以300 mg·L^(-1)最佳;获得转化芽45个,部分株系用PCR和Southern杂交证实外源基因已整合在葡萄基因组DNA中。
Establishing an efficient regeneration and transformation system is a fundamental role to the transgenic study. Using Centennial Seedless as materials, the effects of growth regulators, explants and ablastin to the generation of adventitious shoots were studied and the optimal condition for regeneration and transfor- mation was determined. Results of experiments showed that, when being used alone or combined with auxins, the cytokinin TDZ is always better than BA and KT in inducing adventitious shoots. The adventitious shoots of Centennial Seedless could be efficiently induced in the media of MS ＋ TDZ 2. 0 mg · L^-1, MS ＋ TDZ 4. 0 mg · L^-1 or MS ＋ TDZ 4. 0 mg · L^-1 ＋ IAA 0. 1 mg · L^-1. For fixed medium, adventitious shoots can be generated with minor difference in efficiency from leaf disc, stem and petiole. When this regeneration system was used in genetic transformation mediated by Agrobaterium tumefaciens, the optimal concentration of screening resistance medium was 20 mg · L^-1 for kanamycin, and for the control of agrobacterium growth, the best concentration was 400 mg ·L^-1 for cephaloridine or 300 mg · L^-1 for carbencillin. Forty-five transformants were obtained. It was confirmed by PCR and Southern blotting that the foreign gene had been integrated into the genome of grape.
出处
《园艺学报》
CAS
CSCD
北大核心
2008年第1期27-32,共6页
Acta Horticulturae Sinica
基金
北京市科技新星计划项目(2006B39)
北京市优秀人才培养专项经费资助项目
关键词
葡萄
器官发生途径
不定芽
组织培养
grape
organogenesis approach
adventitious shoot
tissue culture
