A non-antibiotic selection strategy uses the phosphomannose-isomerase (PMI) gene and green fluorescent protein (GFP) gene for Agrobacterium-mediated transformation of Prunus domestica L. leaf explants

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• 作者：Tatiana Sidorova ; Roman Mikhailov…
• 关键词：Fruit trees ; Mannose ; European Plum ; Positive selection ; sgfp ; Genetic transformation
• 刊名：Plant Cell, Tissue and Organ Culture (PCTOC)
• 出版年：2017
• 出版时间：January 2017
• 年：2017
• 卷：128
• 期：1
• 页码：197-209
• 全文大小：
• 刊物类别：Biomedical and Life Sciences
• 刊物主题：Plant Sciences; Plant Physiology; Plant Genetics & Genomics; Plant Pathology;
• 出版者：Springer Netherlands
• ISSN：1573-5044
• 卷排序：128

文摘

We developed an efficient system for agrobacterial transformation of plum (Prunus domestica L.) leaf explants using the PMI/mannose and GFP selection system. The cultivar ‘Startovaya’ was transformed using Agrobacterium tumefaciens strain CBE21 carrying the vector pNOV35SGFP. Leaf explants were placed onto a nutrient medium containing various concentrations and combinations of mannose and sucrose to develop an efficient selection system. Nine independent transgenic lines of plum plants were obtained on a regeneration medium containing 20 g/L sucrose and 15 g/L mannose. The highest transformation frequency (1.40 %) was produced using a delayed selection strategy. Starting from the 1st days after transformation and ending by regeneration of shoots from the transgenic callus, selection of transgenic cells was monitored by GFP fluorescence that allowed avoiding formation of escapes. Integration of the manA and gfp transgenes was confirmed by PCR and Southern blotting. The described transformation protocol using a positive PMI/mannose system is an alternative selection system for production of transgenic plum plants without genes of antibiotic and herbicide resistance, and the use of leaf explants enables retention of cultivar traits of plum plants.