摘要
本研究选用了三个马铃薯(Solanum Iuberosum L.)栽培品种“85T-14-3”、“86-2”及“Favorita”的块茎、微型薯和试管薯为起始材料,应用根癌农杆菌 Ti 质粒系统成功地建立了一种方法简单、速度快和频率高的遗传转化体系。其中试管薯薄片的转化速度最快,经根癌农杆菌(Agrobacterium tumefaciens)共培养后,薄片在100mg/L 卡那霉素的分化培养上,2—3周就可产生出抗性小芽,这些小芽进一步仲长后可在50—100mg/L 卡那霉素的无激素MS 培养基上生根。从共培养到转化植株的获得只需6—7周。微型薯和试管薯的转化频率较高,最高可达67.5%。大多数抗性植株均能检测到胭脂碱合成酶或 GUS 基因的表达。把带有甜蛋白基因和胭脂碱合成酶标记基因的Ti质粒导入马铃薯,获得大量转化植株。叶片抗性检测和 nopaline 检测可推断外源甜蛋白基因已进入马铃薯基因组。
Tuber,minituber and in vitro-grown microtuber discs of potato (Solanum tuberosura L.) cultivars 85—14—3,86—2 and Favorita were used in Agrobacterium mediated gene transfer.A simple,rapid and efficient transformation system was established.Among the three kinds of discs used,the microtuber disc was superior in obtaining transformants.Microtuber discs star- ted to form shoots on shoot inducing medium containing kanamycin two to three weeks after cocultivation.Rooted transformants could be obtained in 6—7 weeks.The transformation effi- ciency could reach as high as 67.5%.The majority of kanamycin resistant plants gave nopaline positive or GUS expression.A number of transgenic plants were obtained using the plasmid containing a sweet protein NPT Ⅱ and nopaline synthase genes.The leaf callus assay and no- paline assay indicated that the foreign sweet protein gene was introduced into the potato genome.
基金
国家重点实验室研究课题
关键词
马铃薯
遗传转化
甜蛋白基因
Potato transformation
Agrobacterium tumefaciens
Sweet protein gene
NPTII
Nopaline systhase
GUS
