杨树无选择标记转基因体系的建立

Establishment of Genetic Transformation System with Selective Marker Free for Poplars

本研究选择美洲黑杨杂种优良无性系南林895杨(Populus×euramericana cv‘.Nanlin895’)为材料,利用Cre/loxP位点特异性重组系统(pX6-GFP载体)进行遗传转化研究。经不同浓度β-雌二醇诱导处理,分别利用农杆菌侵染叶盘分化不定芽至完整植株过程和利用已获得的抗性植株诱导愈伤至植株再生过程删除选择标记基因。结果显示,利用已获得的抗性植株诱导愈伤至植株再生过程删除选择标记基因的效率高于利用农杆菌侵染叶盘分化不定芽至完整植株过程标记基因的删除效率。对卡那霉素的敏感性测试结果表明,已删除选择标记的转基因杨树和对照未转基因南林895杨对卡那霉素的敏感性是一致的。初步建立了杨树无选择标记转基因体系。

Populus×euramericana cv. ‘Nanlin895’, an elite clone of Populus deltoides hybrid was selected as a receptor for genetic transformation by use of Cre/loxP site-specific recombination system （pX6-GFP vector） in this study. The selective markers in the transgenic plants were deleted with different concentration of β-estradiol induction treatment and by use of two methods respectively in one process from differentiation of leaf discs infected by Agrobacterium to whole plants and the other process from callus induced by transgenic plants already obtained to plant regeneration. The results showed that the efficiency of selective marker deletion in the process from callus induced by transgenic plants already obtained to plant regeneration was higher than that in the process from differentiation of leaf discs to whole plants. Results of kanamycin sensitivity test showed that there was the same kanamycin sensitivity between selective marker deleted transgenic plants and the control of Populus×euramericana cv. ‘Nanlin895’. The system of selective marker-free transformation for poplars was initially established in this study.