School of Biological Sciences, The University of Hong Kong, Pokfulam, Hong Kong, China.
J Agric Food Chem. 2013 Jul 3;61(26):6423-9. doi: 10.1021/jf401537q. Epub 2013 Jun 25.
Plants are proven effective bioreactors for the production of heterologous proteins including those desired by the biopharmaceutical industry. However, the potential of plants as bioreactors is limited by the availability of characterized plant promoters that can drive target gene expression in relatively distant plant species. Seeds are ideal for protein storage because seed proteins can be kept stably for several months. Hence, a strong promoter that can direct the expression and accumulation of target proteins within seeds represents a powerful tool in plant biotechnology. Toward this end, an effort was made to identify such a promoter from Vigna radiata (mung bean) to drive expression in dicot seeds. A 784-bp 5'-flanking sequence of the gene encoding the 8S globulin α' subunit (8SGα') of the V. radiata seed storage protein was isolated by genome walking. When the 5'-flanking region was analyzed with bioinformatics tools, numerous putative cis-elements were identified. The Green Fluorescent Protein (GFP) regulated by this promoter was observed to be transiently expressed in protoplasts derived from V. radiata cotyledons. Finally, transgenic Arabidopsis plants expressing the β-glucuronidase (GUS) reporter gene driven from the 8S globulin α' promoter showed strong GUS expression in transgenic embryos in both histochemical and quantitative GUS assays, confirming high expression within seeds. Therefore, the V. radiata 8S α' promoter has shown potential in directing expression in seeds for bioreactor applications.
植物已被证实是生产异源蛋白的有效生物反应器,包括那些生物制药行业所需的蛋白。然而,植物作为生物反应器的潜力受到可用的已鉴定植物启动子的限制,这些启动子可以在相对较远的植物物种中驱动靶基因的表达。种子是蛋白质储存的理想选择,因为种子蛋白可以稳定保存数月。因此,能够在种子内指导目标蛋白表达和积累的强启动子是植物生物技术中的有力工具。为此,人们努力从豇豆(绿豆)中鉴定出这样一个启动子,以在双子叶种子中驱动表达。通过基因组步移,分离到编码 Vigna radiata 种子贮藏蛋白 8S 球蛋白α'亚基(8SGα')的基因的 784bp 5'-侧翼序列。当用生物信息学工具分析 5'-侧翼区时,鉴定出了许多假定的顺式元件。受该启动子调控的绿色荧光蛋白(GFP)在 V. radiata 子叶原生质体中瞬时表达。最后,表达由 8S 球蛋白α'启动子驱动的β-葡萄糖醛酸酶(GUS)报告基因的转基因拟南芥植物在组织化学和定量 GUS 分析中显示出转基因胚胎中强烈的 GUS 表达,证实了种子内的高表达。因此,豇豆 8Sα'启动子在指导生物反应器应用中的种子表达方面显示出了潜力。