Division of Biology, Faculty of Natural Sciences, Imperial College London, London, UK.
Plant Biotechnol J. 2011 Jun;9(5):575-84. doi: 10.1111/j.1467-7652.2010.00575.x. Epub 2010 Nov 24.
Chloroplast transformation is a promising approach for the commercial production of recombinant proteins in plants. However, gene containment still remains an issue for the large-scale cultivation of transplastomic plants in the field. Here, we have evaluated the potential of using tobacco transplastomic cell suspensions for the fully contained production of a modified form of the green fluorescent protein (GFP+) and, a vaccine antigen, fragment C of tetanus toxin (TetC). Expression of these proteins in cell suspension cultures (and calli) was much less than in leaves, reaching 0.5%-1.5% of total soluble protein (TSP), but still produced 2.4-7.2 mg/L of liquid culture. Much better expression levels were achieved with a novel protein production platform in which transgenic cell suspension cultures were placed in a temporary immersion bioreactor in the presence of Thidiazuron to initiate shoot formation. GFP+ yield reached 660 mg/L of bioreactor (33% TSP), and TetC accumulated to about 95 mg/L (8% TSP). This new production platform, combining the rapid generation of transplastomic cell suspension cultures and the use of temporary immersion bioreactors, is a promising route for the fully contained low-cost production of recombinant proteins in chloroplasts.
叶绿体转化是在植物中商业化生产重组蛋白的一种很有前途的方法。然而，对于在田间大规模种植转基因叶绿体植物来说，基因控制仍然是一个问题。在这里，我们评估了利用烟草转基因叶绿体悬浮细胞进行完全封闭生产改良型绿色荧光蛋白（GFP+）和破伤风毒素片段 C（TetC）疫苗抗原的潜力。这些蛋白在悬浮细胞培养物（和愈伤组织）中的表达量远低于叶片，仅达到总可溶性蛋白（TSP）的 0.5%-1.5%，但液体培养物仍可产生 2.4-7.2mg/L。通过一种新的蛋白生产平台，可实现更好的表达水平，该平台中，在存在噻唑隆的情况下，将转基因悬浮细胞培养物置于临时浸没生物反应器中，以启动芽的形成。GFP+的产量达到生物反应器的 660mg/L（33%TSP），而 TetC 积累到约 95mg/L（8%TSP）。这种新的生产平台，结合了转基因叶绿体悬浮细胞培养物的快速生成和临时浸没生物反应器的使用，是在叶绿体中进行完全封闭、低成本生产重组蛋白的一种很有前途的途径。
