Single-vector three-frame expression systems for affinity-tagged proteins

摘要: An effort is presented to create expression vectors which would allow of an inserted gene fragment in three reading frames a single vector from promoter but with separate ribosome binding sites (RBS). Each frame generate in-frame fusion affinity tag efficient recovery the produced proteins. In first generation vector, identical polyhistidyl tags (His6) were used as for frames. second different tags, albumin domain derived streptococcal protein G, IgG Staphylococcus aureus A-derived (Z) and His6 tag, employed frame-specific recovery. To evaluate systems, model genes have been both vectors. The was demonstrated produce proteins all frames, whereas second, much wider spacing between RBSs could only be two translational start sites. For translation found significantly favored over others. Nevertheless, we believe that results represent successful attempt single-vector three-frame concept become valuable future combined cloning–expression