Transformation of Upland Cotton (Gossypium hirsutum L.) with gfp Gene as a Visual Marker
Transformation of Upland Cotton (Gossypium hirsutum L.) with gfp Gene as a Visual Marker
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以 gfp 基因作为视觉标记转化陆地棉 (Gossypium hirsutum L.)
DOI：
10.1016/s2095-3119(12)60081-1
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发表时间：
2012-01-01
影响因子：
4.8
通讯作者：
Zhang Xian-long
中科院分区：
文献类型：
--
作者：
Jin Shang-xia;Liu Guan-ze;Zhang Xian-long
将绿色荧光蛋白(GFP)基因作为棉花(Gossypium hirsutom L.)转化和植物再生。转基因细胞在与农杆菌共培养42h后即可观察到绿色荧光蛋白的高表达。大多数稳定转化事件发生在原生维管组织的细胞中。外植体共培养4d后均可检测到GFP的瞬时表达，但共培养3d时GFP的稳定表达最高。我们认为外源基因在遗传转化中的瞬时表达和稳定表达是两个相对但不同的事件，因为高瞬时表达并不一定会导致高稳定的转化。在相同的体外培养条件下，转基因和非转基因愈伤组织在不同发育阶段表现出不同的形态特征。高浓度的植物生长调节剂(PGRs)对转基因愈伤组织的体细胞胚胎发生是有效的，这意味着转基因愈伤组织比非转基因愈伤组织进一步生长和体细胞胚胎发生所需的激素剂量相对较高。在T_1代的叶、茎、叶柄、花组织和幼苗中均有较强的GFP表达。对8个转基因株系在T_1代中表达GFP的分离比进行了评分，进一步证明了转基因植株的稳定转化。这些结果证明了GFP是一个在整个转化事件中对协议进行优化、选择和监测的强大报告基因。
The green-fluorescent protein (gfp) gene was evaluated as a screening marker during cotton (Gossypium hirsutum L.) transforming and plant regeneration. High expression of GFP (green-fluorescent protein) was observed in transgenic cells as early as 42 h after co-culture with Agrobacterium. Most of the stable transformation events were detected in the cells of primary vascular tissue. GFP transient expression could be detected on all the explants after co-culturing for 4 d, however, the highest GFP stable expression was recorded when the explants were co-cultured for 3 d. We believe the transient and stable expression of a foreign gene in genetic transformation were two relative but different events, because high transient expression did not surely lead to high stable transformation. Under the same conditions of in vitro culture, transgenic and non-transgenic calli exhibited different morphological characters on different stages of development. High concentration of plant growth regulators (PGRs) was efficient for somatic embryogenesis of the transgenic calli, which means that the transgenic calli need relatively higher dose of hormone for further growth and somatic embryogenesis than non-transgenic ones. Strong GFP-expression was observed in leaf, stem, petioles, floral tissues, and seedlings of T, progeny. Segregation ratios of eight transgenic lines were scored for expression of GFP in the T, progeny that providing further evidence of stable transformation. These results proved that GFP is a powerful reporter gene for protocol optimization, selection, and monitioring in whole transformation events.