Oker-Blom C, Orellana A, Keinanen K
VTT Biotechnology and Food Research, Espoo, Finland.
FEBS Lett. 1996 Jul 8;389(3):238-43. doi: 10.1016/0014-5793(96)00593-5.
We have generated recombinant baculoviruses for expression of the green fluorescent protein (GFP), a bright GFP mutant (S65T), and a GFP-streptavidin fusion protein in Sf9 and High Five insect cell lines. At 3-4 days post infection, about 30% of the total protein contents was represented by the recombinant protein products, giving the infected insect cells a bright green color which was clearly visible by eye in daylight. The isolated GFP-streptavidin fusion protein, which possessed fluorescence properties identical to those of the native GFP, was capable of binding biotin as shown by using biotinylated beads as well as biotinylated antibody complexes decorating surface expressed GluR-6 glutamate receptor in live and fixed insect cells. The exceptionally high expression levels of GFP and GFP (S65T) and the GFP-streptavidin fusion protein in recombinant baculovirus infected insects should facilitate production of GFP derivatives for in vitro applications.
我们已经构建了重组杆状病毒，用于在Sf9和High Five昆虫细胞系中表达绿色荧光蛋白（GFP）、一种明亮的GFP突变体（S65T）以及一种GFP-链霉亲和素融合蛋白。在感染后3 - 4天，重组蛋白产物占总蛋白含量的约30%，使被感染的昆虫细胞呈现明亮的绿色，在日光下肉眼清晰可见。分离出的GFP-链霉亲和素融合蛋白具有与天然GFP相同的荧光特性，通过使用生物素化磁珠以及在活的和固定的昆虫细胞中用生物素化抗体复合物修饰表面表达的GluR-6谷氨酸受体，表明其能够结合生物素。重组杆状病毒感染昆虫中GFP、GFP（S65T）和GFP-链霉亲和素融合蛋白的极高表达水平应有助于生产用于体外应用的GFP衍生物。
