Epicotyl sections as targets for plant regeneration and transient transformation of common bean using Agrobacterium tumefaciens

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• 作者：R. Collado ; I. Bermúdez-Caraballoso…
• 关键词：Agrobacterium tumefaciens ; Genetic transformation ; GUS activity ; Phaseolus vulgaris ; Epicotyl section
• 刊名：In Vitro Cellular & Developmental Biology - Plant
• 出版年：2016
• 出版时间：October 2016
• 年：2016
• 卷：52
• 期：5
• 页码：500-511
• 全文大小：1,008 KB
• 刊物主题：Plant Sciences; Cell Biology; Developmental Biology; Plant Breeding/Biotechnology; Plant Genetics & Genomics;
• 出版者：Springer US
• ISSN：1475-2689
• 卷排序：52

文摘

Common bean is recalcitrant to genetic transformation, due to limited regeneration capacity and low DNA transfer rates. The effect of different parameters on T-DNA transfer from Agrobacterium tumefaciens, was studied by measuring transient expression of the β-glucuronidase gene in Phaseolus vulgaris L. cv. CIAP7247F. Epicotyl containing seedling explants were inoculated with Agrobacterium EHA101 and C58C1RifR(pMP90) strains harboring the binary vector pTJK136 with GusA gene on the T-DNA. Parameters studied were temperature and light regime during co-cultivation, explant injury, and the acetosyringone concentration for vir gene induction. The co-cultivation temperature and photoperiod had a significant effect on Agrobacterium DNA transfer. In addition, explant injury and supplementation of the co-cultivation medium with acetosyringone increased the GUS activity. Optimal T-DNA transfer was obtained under the following conditions: co-cultivation at 25°C in darkness, injuring the explants with carborundum, and supplementation of the co-cultivation medium with 200 μM acetosyringone. This T-DNA delivery system was combined with a direct organogenesis protocol using epicotyl explants and fertile regenerants were recovered from tissue transformed with Agrobacterium. However, no transmission of transgenes to progeny could be observed, suggesting that the obtained plants were chimeras.