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首页> 外文期刊>In Vitro Cellular and Development Biology. Plant: Journal of the Tissue Culture Association >Factors enhancing Agrobacterium tumefaciens-mediated gene transfer in peanut (Arachis hypogaea L.).
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Factors enhancing Agrobacterium tumefaciens-mediated gene transfer in peanut (Arachis hypogaea L.).

机译：增强根癌农杆菌介导的花生中基因转移的因子。

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摘要

Parameters enhancing Agrobacterium-mediated transfer of foreign genes to groundnut (Arachis hypogaea) cells were investigated. An intron-containing beta-glucuronidase uidA (gusA) gene under the transcriptional control of CaMV 35S promoter served asa reporter. Transformation frequency was evaluated by scoring the number of sectors expressing GUS activity on leaf and epicotyl explants. The 'Valencia Select' market type cv. New Mexico was more amenable to Agrobacterium transformation than the 'runner' market type cultivars tested (Florunner, Georgia Runner, Sunrunner, or South Runner). The disarmed Agrobacterium tumefaciens strain EHA101 was superior in facilitating the transfer of uidA gene to groundnut cells compared to the disarmed strain C58. Rinsing of explants in half-strength Murashige-Skoog (MS) media prior to infection by Agrobacterium significantly increased the transformation efficiency. The use of cocultivation media containing high auxin (1.0 or 2.5 mg/litre (4.53 鍹 or 11.31 鍹) 2,4-D) and low cytokinin (0.25 or 0.5 mg/litre (1.0 鍹 or 2.0 鍹) BA) promoted higher rates of transformation than either hormone-free or thidiazuron-containing medium. The polarity of the epicotyl during cocultivation was important; explants incubated in aninverted (vertically) manner followed by a vertically upright position resulted in improved transformation and shoot regeneration frequencies. Preculture of explants in MS basal medium or with 2.5 mg thidiazuron/litre prior to infection markedly decreased the number of transformed zones. The optimized protocol was used to obtain transient transformation frequencies ranging from 12% to 36% for leaf explants and 15% to 42% for epicotyls. Initial evidence of transformation was obtained by polymerase chainreaction and subsequently confirmed by Southern analysis of regenerated plants.
机译：研究了增强农杆菌介导的外源基因向花生（花生）的转移的参数。在CaMV 35S启动子的转录控制下，含内含子的β-葡萄糖醛酸酶uidA（gusA）基因作为报告基因。通过对在叶和表皮外植体上表达GUS活性的区段的数量进行评分来评估转化频率。 “ Valencia Select”市场类型cv。新墨西哥州比经过测试的“跑步者”市场类型的品种（Florunner，Georgia Runner，Sunrunner或South Runner）更适合农杆菌转化。与解除武装的农杆菌菌株C58相比，解除武装的土壤杆菌农杆菌菌株EHA101在促进uidA基因向花生细胞转移方面具有优势。在农杆菌感染之前，在半强度Murashige-Skoog（MS）培养基中冲洗外植体可显着提高转化效率。使用含有高生长素（1.0或2.5毫克/升（4.53鍹或11.31鍹）2,4-D）和低细胞分裂素（0.25或0.5毫克/升（1.0鍹或2.0 BA）BA的共培养培养基可以提高收率转化率要高于不含激素或含噻唑隆的培养基。共培养过程中上胚轴的极性很重要。以反向（垂直）方式温育的外植体，然后垂直竖立放置，可提高转化和芽再生的频率。在感染前在MS基础培养基中或以2.5 mg噻唑隆/升对外植体进行预培养可显着减少转化区的数量。优化的方案用于获得瞬时转化频率，其中外植体的瞬时转化频率范围为12％至36％，而上胚轴的瞬时转化频率范围为15％至42％。转化的初步证据是通过聚合酶链反应获得的，随后通过对再生植物的Southern分析得到了证实。

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