Yanhua Zhao

Hebei Academy of Agricultural and Forestry Sciences, Shih-chia-chuang, Shanxi Sheng, China

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Publications (2)1.5 Total impact

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    ABSTRACT: 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.Highlights► Wounding treatment promotes adventitious shoot regeneration from fragmented shoot tips. ► Cells of fragmented shoot tips were more resistant to kanamycin selection compared to leaf cells. ► Transgenic apple plants were obtained from fragmented shoot tips. ► Shoot regeneration from transgenic cells was a major factor inhibiting transformation efficiency.
    Scientia Horticulturae 01/2011; 128(4):450-456. · 1.50 Impact Factor
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    ABSTRACT: Non-forest woody plants include many fruit trees and ornamental plants. They not only provide delicious fruit but are also important as horticultural landscape plants. Conservation of these germplasm resources is necessary for the continued development of agriculture. As most species and cultivars of woody plants are genetically heterozygous, their genetic integrity must be maintained through vegetative propagation. The conventional method for woody plants is conservation in the field. However for longterm preservation of these genetic resources the field genebank is not only costly, because of the land and constant maintenance, but also susceptible to insects, diseases and environmental stress (Engelmann 2000; Reed et al. 2005). In-vitro cultured plantlets can be mass propagated with limited space and under disease-free conditions. Most cultures can be held under reduced or minimum growth conditions for years without reculture. Thus in-vitro plantlets are suitable materials for short- or medium-term backup storage of plant germplasm (Engelmann 2003; Reed and Chang 1997). Cryopreserved storage of plants in liquid nitrogen (LN) is the most promising approach to achieve long-term maintenance of woody plant germplasm (Sakai 1995). Cryopreservation can save labor and space, and it complements current germplasm storage methods, making it an important tool for long-term storage of vegetatively propagated plants (Engelmann 2000). Sakai (1960) first demonstrated that winter-hardy twigs survived after immersion in LN for 1 year. Since then many reports indicate that materials with meristems or embryogenic cultures of woody plants can be conserved successfully in LN (Kartha 1985, Sakai 1995, Reed and Chang 1997).
    12/2007: pages 387-420; Springer Science and Business Media LLC.