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作 者:曹冬梅[1] 康黎芳[1] 李永平[1] 段九菊[1] 张超[1] 王云山[1]
机构地区:[1]山西省农业科学院园艺研究所,太原030031
出 处:《农业生物技术学报》2015年第3期329-336,共8页Journal of Agricultural Biotechnology
基 金:山西省回国留学人员科研资助项目(No.2010-117);山西省农科院育种基础项目(No.yyzjc0925)
摘 要:安祖花(Anthurium andraeanum)是近年来非常流行的年宵花卉之一.为了优化安祖花的遗传转化体系,本研究以安祖花Arizona品种组培苗的叶片为外植体,将构建好的重组质粒pSN 1301-HYG-PhCHS在根癌农杆菌(Agrobacterium tumefaciens) GV3101介导下转化安祖花.优化后的体系为:首先将组培苗幼嫩叶片切块后置于1/3MS预培养基(附加200 mg/L NH4NO3,0.5 mg/L BA,0.05 mg/L 2,4-D,5 g/L琼脂,30 g/L蔗糖)中预培育3d,然后用根癌农杆菌(菌液OD600=0.5)侵染10 min,将侵染过的叶片接种于MS培养基,28℃黑暗条件下共培养3d,之后转移到附加有500 mg/L羧苄青霉素和30 mg/L潮霉素的愈伤诱导培养基中进行筛选.结果表明,抗性愈伤组织形成率可达到5%~6%.对体系优化过程中获得的14株抗性转基因植株进行PCR检测,其中9株为阳性,证实矮牵牛查尔酮合成酶基因(Petunia hybrida chalcone synthase gene,PhCHS)已经转入安祖花Arizona中.该研究结果为利用转基因方法改良安祖花品种提供了技术保障.Anthurium andraeanum is one of the popular lunar new year flowers in recent years. In order to optimize genetic transformation system for Anthurium andraeanum, leaves of cultivar Arizona plantlets was used as explants and the recombinant plasmid pSN1301-HYG-PhCHS was transferred to A. andraeanum cv. Arizona mediated by Agrobacterium tumefaciens GV3101. The efficient transformation system for A. andraeanum cultivar Arizona was described as follows: Leaves were cut and pre-cultured for 3 d on the 1/3 MS culture medium, which contained 200 mg/L NI-LNO3, 0.5 mg/L BA, 0.05 mg/L 2,4-D, 5 g/L Agar and 30 g/L Sucrose, and then infected with A. tumefaciens (0D6oo=0.5) for 10 min. The inoculated leaves were put on MS culture medium at 28 ~C under dark conditions for 3 d, and then transferred to the callus-inducing medium contained 30 mg/L hygromycin for resistant screening. The percentage of resistant-calli was 5%-6%. In total 14 transgenic plants with hygromycin resistance were obtained in this study and detected by PCR, and 9 of them were verified to be positive transgenic plants. PCR analysis also confirmed that Petunia hybrida chalcone synthase gene (PhCHS) has been transferred into A. andraeanum cultivar Arizona. The results will provide the technical support for the improvement of transgenic method forAnthurium cultivars.
关 键 词:安祖花 遗传转化 抗性愈伤组织形成率 矮牵牛查尔酮合成酶基因(PhCHS)
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