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The effects of plasmid content, transcription efficiency, and translation efficiency on the productivity of a cloned gene protein in Escherichia coli
Author(s) -
Kim JeongYoon,
Ryu Dewey D. Y.
Publication year - 1991
Publication title -
biotechnology and bioengineering
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.136
H-Index - 189
eISSN - 1097-0290
pISSN - 0006-3592
DOI - 10.1002/bit.260381103
Subject(s) - plasmid , pbr322 , biology , gene , microbiology and biotechnology , transcription (linguistics) , translational efficiency , gene expression , expression vector , escherichia coli , promoter , heterologous expression , protein biosynthesis , lac operon , heterologous , recombinant dna , translation (biology) , genetics , messenger rna , linguistics , philosophy
In order to investigate how plasmid content, transcription efficiency, and translation efficiency affect the productivity of a cloned gene protein, a new vector (pPLc‐RP4.5) was constructed. The vector has P L promoter and lacZ as a structure gene 4.5S RNA gene between P L promoter and lacZ gene. We took advantage of the characteristic that the 4.5S RNA is accumulated inside E. coli cells and can be quantitatively measured. A two‐stage continuous culture system in combination with a temperature‐sensitive gene switching system was used to study the performance of the recombinant fermentation. It was found that the plasmid content as varied by the dilution rate in the production stage showed a different pattern from that in the growth stage. The result showed that promoter strength had a greater influence on the overall gene expression efficiency of a cloned gene than the plasmid content, and the overall gene expression efficiency was largely dependent upon translation efficiency when a multicopy plasmid (pBR322 derivative and rop ‐) and a strong promoter (P L ) were used to express a heterologous protein in E. coli .