Use of the human elongation factor 1 alpha promoter as a versatile and efficient expression system.
We have characterized the promoter region of the human elongation factor 1 alpha-encoding gene (EF-1 alpha) and developed a versatile expression system which has a wide host range and a high efficiency of gene expression. To identify the promoter region of the EF-1 alpha gene necessary for efficient gene expression, we constructed four pEF-CAT plasmids that have the bacterial cat gene fused to four different sites of the human EF-1 alpha gene: (i) ligated to the end of the TATA box (pEF220-CAT); (ii) ligated in exon 1 (pEF204-CAT and pEF233-CAT), and (iii) ligated in exon 2 (pEF321-CAT). All the pEF-CAT plasmids were highly expressed in all the cell types tested, including fibroblasts and lymphoid cells. Plasmid pEF321-CAT, which contains the first exon and the first intron, gave the highest level of cat expression. Plasmids pEF204- and pEF233-CAT, which contain part of the first exon but do not contain the first intron, were less efficient in cat expression than was pEF321-CAT. Plasmid pEF220-CAT, which lacks both the first exon and the first intron, was the least efficient. Plasmid pEF321-CAT was several- to 100-fold more efficient in cat expression than plasmid pSV2-CAT depending on the recipient cell types. The promoter of pEF321 plasmid also directed the stable expression of the bacterial neo gene more efficiently than the promoter of the simian virus 40 (SV40) early gene or the long terminal repeat of Rous sarcoma virus. Using this system, the SV40 early gene and the cDNA encoding human CD4 were also expressed efficiently.
Department of Virology, Institute of Medical Science, University of Tokyo, Japan.
D W
DW
Kim
T
T
Uetsuki
Y
Y
Kaziro
N
N
Yamaguchi
S
S
Sugano
D000818:Animals; D000957:Antigens, Viral, Tumor / Q000235:genetics; D015704:CD4 Antigens / Q000235:genetics; D002460:Cell Line; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics / Q000378:metabolism; D003001:Cloning, Molecular; D005091:Exons; D005786:Gene Expression Regulation; D005822:Genetic Vectors; D006801:Humans; D007438:Introns; D020648:Peptide Elongation Factor 1; D010445:Peptide Elongation Factors / Q000235:genetics*; D010957:Plasmids; D011401:Promoter Regions, Genetic; D015183:Restriction Mapping; D014162:Transfection; D014407:Tumor Cells, Cultured
D000957:Antigens, Viral, Tumor; D015704:CD4 Antigens; D020648:Peptide Elongation Factor 1; D010445:Peptide Elongation Factors; D015500:Chloramphenicol O-Acetyltransferase