Tac promoter vectors incorporating the bacteriophage T7 gene 10 translational enhancer sequence for improved expression of cloned genes in Escherichia coli.
Two new plasmid expression vectors, pTacT7 and pTacT7L, have been constructed, which incorporate between the tac promoter and a downstream NcoI-HindIII polylinker sequence a synthetic sequence derived from the region upstream from gene 10 of bacteriophage T7 (g10-L). This sequence was recently shown to act as a translational enhancer (Olins et al., 1988) and was termed "Epsilon" (Enhancer of Protein Synthesis Initiation) element (Olins and Rangwala, 1989). In this communication we describe in detail the construction of ptacT7 and ptacT7L. Furthermore, we present evidence that the "Epsilon" element is able to enhance 3 to 20-fold the expression levels of two poorly expressed test genes encoding the human placental proteins PP9 and PP15. On the other hand, the expression levels of two highly expressed test genes encoding the human placental proteins PP4 and FXIIIa could not be further enhanced by the presence of the "Epsilon" element. These experiments show that the T7 gene 10 leader sequence can be utilized to improve the expression yields of otherwise poorly expressed heterologous genes in Escherichia coli.
Research Laboratories of Behringwerke AG, Marburg, F.R.G.
B
B
Lehmeier
E
E
Amann
D001483:Base Sequence; D015153:Blotting, Western; D003001:Cloning, Molecular / Q000379:methods*; D004279:DNA, Viral; D004742:Enhancer Elements, Genetic; D004926:Escherichia coli / Q000235:genetics*; D005822:Genetic Vectors; D006801:Humans; D008969:Molecular Sequence Data; D011257:Pregnancy Proteins / Q000235:genetics; D011401:Promoter Regions, Genetic; D014176:Protein Biosynthesis; D013604:T-Phages / Q000235:genetics*
D004279:DNA, Viral; D011257:Pregnancy Proteins