渐狭叶烟草组培苗生根条件优化及在基因转化中的应用

针对渐狭叶烟草(Nicotiana attenuata)组培苗生根困难的问题，本研究测试了不同浓度萘乙酸(NAA)、吲哚丁酸(IBA)、吲哚乙酸(IAA)、嘧啶醇、矮壮素、生根粉及活性炭等外源添加物对组培苗生根的影响。结果表明，对照处理的组培苗在90 d内基本不生根；不同浓度矮壮素处理也不能诱导组培苗生根；而合适浓度的NAA、IBA或IAA处理均能较好地诱导组培苗生根，其中0.8 mg·L^-1 IBA不仅具有较好的诱导生根效果，生根率达72.73%，且根及地上部分均生长良好。进一步测试发现0.8 mg·L^-1 IBA处理对转基因组培苗具有良好的生根效果，20 d后生根率达60%以上，而对照组未生根；经伤口诱导和0.8 mg·L^-1 IBA处理，40 d内总生根率可达90%以上。该研究为高效获得渐狭叶烟草转基因生根苗提供了解决方案，也为渐狭叶烟草作为烟草属模式植物的研究提供了便利。

Optimization of Rooting Conditions in Tissue-cultured Seedlings of Nicotiana attenuata and Its Applications

Nicotiana attenuata, a diploid wild tobacco, has recently became the most important model plant of Nicotiana species for studying plant-herbivore and plant-pathogen interaction, as it has relative smaller genome and shorter life cycle when compared to cultivated tobacco. However, the difficulty of rooting during tissue culture of transgenic N. attenuata seedlings greatly hampered its application as a research model in genetic engineering and molecular biology research. In order to overcome this difficulty, we tested the effects of exogenous additives, such as naphthylacetic acid (NAA), indole-3-butyric acid (IBA), indole-3-acetic acid (IAA), ancymidol, chlormequat chloride (CCC), rooting powder and active charcoal with various concentrations, on the rooting of tissue-cultured seedlings. The results revealed that the seedlings hardly produced any roots without any treatments or with CCC treatments during growth in rooting medium for 90 days, while treatments with NAA, IBA, IAA or rooting powder with appropriate concentration greatly induced rooting of tissue-cultured seedlings; Seedlings in the rooting medium supplied with 0.8 mg·L^-1 IBA not only achieved a good rooting rate with above 72.73%, but also grew well in terms of morphology of roots and aboveground parts. Finally, we found that 0.8 mg·L^-1 IBA treatments also greatly increased the rooting rate of transgenic seedlings, from 0% (control) to more than 60% after 20 days in rooting medium, and reached to more than 90% after wounding treatments and sub-culturing in rooting medium supplied with 0.8 mg·L^-1 IBA for another 20 days. Taken all together, we successfully optimized the rooting protocol for N. attenuata transformation. The findings of this study provided a solution for the efficient induction of roots in transgenic seedlings of N. attenuata, and brought great convenience for using N. attenuata as a model plant of tobacco species.