文献类型： 外文期刊

作者： Wu, Yongjie ¹ ; Li, Yunhe ² ; Wu, Yaqin ¹ ; Cheng, Hehe ¹ ; Li, Yin ³ ; Zhao, Yanhua ¹ ; Li, Yusheng ¹ ;

作者机构： 1.Hebei Acad Agr & Forestry Sci, Changli Inst Pomol, Changli Town 066600, Peoples R China

2.Chinese Acad Trop Agr Sci, S Subtrop Crop Res Inst, Zhanjiang 524091, Peoples R China

3.Sun Yat Sen Univ, Sch Life Sci, Guangzhou 510275, Guangdong, Peoples R China

关键词： kanamycin: 8063-07-8;green fluorescent protein: GFP;expression;callus formation;adventitious shoot regeneration;genetic breeding

期刊名称：SCIENTIA HORTICULTURAE （ 影响因子：3.463； 五年影响因子：3.672 ）

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收录情况： SCI

摘要： Transgenic apple (Malus baccata ( L) Borkhausen) plants were obtained via Agrobaterium-mediated transformation of fragmented shoot tips. Our results showed that without wounding treatment or with wounding treatment (II, cutting shoot tips vertically into two parts), shoots generally regenerated from meristem tissues directly and adventitious shoot regeneration was rarely observed. Otherwise, when shoot tips were cut vertically into four parts, a high percentage of callus formation (89.2%) and of adventitious shoot regeneration (60.8%) was observed. Under 20 mg l(-1) kanamycin selection pressure, over 51.7% fragmented shoot tips developed callus and seven transgenic plants with GFP (Green fluorescent protein) expression were obtained from about 120 explants (efficiency of 5.8%). No transgenic plant was obtained from agrobacteria mediate transformed leaves, even though 23.2% of which formed callus after co-cultivation and selection. Molecular analysis (PCR and RT-PCR) of the transformed lines with GFP expression confirmed integration and transcription of the transgene. Under fluorescence microscopy, areas with high density of transgenic cells were observed at the cutting edges of fragmented shoot tips, which indicated that shoot regeneration from transgenic cells should be a major factor inhibiting transformation efficiency. Our experiments also showed that with moderate or low selection pressure, transgenic shoots were obtained generally accompanied by a high numbers of chimeric shoots. While by using fluorescence microscopy observation of GFP expression, the transgenic and chimeric shoots could be detected and separated precisely for further transgenic plats regeneration or multiplication. This may be very useful for apple genetic breeding, as large numbers of transgenic plants could be obtained in a short time