Henrique-Silva Flavio, Soares-Costa Andrea
Laboratory of Molecular Biology, Department of Genetic and Evolution, Federal University of São Carlos, São Carlos, SP, Brazil.
Methods Mol Biol. 2012;847:437-50. doi: 10.1007/978-1-61779-558-9_34.
Transgenic plants have been widely used as expression systems of recombinant proteins in recent years because it can be an efficient alternative for the large-scale production of proteins. This is an area with great potential but is still not much explored. Indeed, this system can bring a breakthrough in the expression of any protein. The model used here as a protein factory was sugarcane, a crop of great global importance. This chapter describes the system that has been adopted in the routine production of transgenic sugarcane coupled with protein purification protocol. In this chapter, we describe production of transgenic sugarcane expressing a His-tagged cystatin under the control of the maize ubiquitin promoter. A transformed sugarcane plant presented high levels of protein expression and was selected for the purification of this protein through affinity chromatography in a nickel column. These studies demonstrate that sugarcane can be a viable expression system for recombinant protein production and that the His-tag purification strategy used to isolate the purified protein was effective.
近年来,转基因植物已被广泛用作重组蛋白的表达系统,因为它可以成为大规模生产蛋白质的有效替代方法。这是一个具有巨大潜力但仍未得到充分探索的领域。事实上,这个系统可以在任何蛋白质的表达上带来突破。这里用作蛋白质工厂的模型是甘蔗,一种具有全球重要性的作物。本章描述了在转基因甘蔗的常规生产中采用的系统以及蛋白质纯化方案。在本章中,我们描述了在玉米泛素启动子控制下表达带有His标签的半胱氨酸蛋白酶抑制剂的转基因甘蔗的生产。一株转化的甘蔗植株呈现出高水平的蛋白质表达,并被选择通过镍柱亲和色谱法纯化该蛋白质。这些研究表明,甘蔗可以成为重组蛋白生产的可行表达系统,并且用于分离纯化蛋白的His标签纯化策略是有效的。