Valkov Vladimir T, Gargano Daniela, Scotti Nunzia, Cardi Teodoro
CNR-IBBR, National Research Council of Italy, Institute of Biosciences and BioResources, Naples, Italy.
Methods Mol Biol. 2014;1132:295-303. doi: 10.1007/978-1-62703-995-6_18.
Although plastid transformation has attractive advantages and potential applications in plant biotechnology, for long time it has been highly efficient only in tobacco. The lack of efficient selection and regeneration protocols and, for some species, the inefficient recombination using heterologous flanking regions in transformation vectors prevented the extension of the technology to major crops. However, the availability of this technology for species other than tobacco could offer new possibilities in plant breeding, such as resistance management or improvement of nutritional value, with no or limited environmental concerns. Herein we describe an efficient plastid transformation protocol for potato (Solanum tuberosum subsp. tuberosum). By optimizing the tissue culture system and using transformation vectors carrying homologous potato flanking sequences, we obtained up to one transplastomic shoot per bombardment. Such efficiency is comparable to that usually achieved in tobacco. The method described in this chapter can be used to regenerate potato transplastomic plants expressing recombinant proteins in chloroplasts as well as in amyloplasts.
尽管质体转化在植物生物技术中具有诱人的优势和潜在应用,但长期以来它仅在烟草中具有高效性。缺乏高效的筛选和再生方案,并且对于某些物种而言,在转化载体中使用异源侧翼区域进行重组效率低下,这阻碍了该技术向主要作物的扩展。然而,除烟草外的其他物种能够利用这项技术,这可能为植物育种提供新的可能性,例如抗性管理或营养价值的改善,且对环境的影响很小或没有影响。在此,我们描述了一种针对马铃薯(Solanum tuberosum subsp. tuberosum)的高效质体转化方案。通过优化组织培养系统并使用携带同源马铃薯侧翼序列的转化载体,我们每次轰击可获得多达一个转基因质体芽。这种效率与烟草中通常实现的效率相当。本章所述方法可用于再生在叶绿体以及造粉体中表达重组蛋白的马铃薯转基因质体植物。