Construction of a versatile promoter analysis vector and its use for analysis of the Serratia marcescens aspartase promoter region.
A new versatile promoter analysis vector, pLGlacZ7, which contains a multiple cloning site and the lacZ structural gene in a low-copy-number plasmid pLG339, has been constructed. This plasmid, which can be stably maintained in Escherichia coli and Serratia marcescens, is useful for analysis of gene expression using lacZ gene fusions. The multiple cloning site of pLGlacZ7 is convenient for the insertion or deletion of promoter DNA fragments, the latter by using exonuclease III. The promoter of the S. marcescens aspA gene encoding aspartase was analyzed using plasmid pLGlacZ7. The S. marcescens aspA gene is composed of 1434 nucleotides and codes for a protein with a Mr of 52,543 whose predicted amino acid sequence was very similar to that of the E. coli aspA gene product. Two functional regions that may participate in the transcription of the S. marcescens aspA gene were found in the promoter region by using lacZ gene fusions in pLGlacZ7.
Omori
K
K
Research Laboratory of Applied Biochemistry, Tanabe Seiyaku Company Limited, Osaka, Japan.
Akatsuka
H
H
Komatsubara
S
S
D000595:Amino Acid Sequence; D001220:Aspartate Ammonia-Lyase / Q000235:genetics*; D001426:Bacterial Proteins / Q000235:genetics*; D001483:Base Sequence; D003001:Cloning, Molecular; D004262:DNA Restriction Enzymes; D004269:DNA, Bacterial / Q000235:genetics*; D005798:Genes, Bacterial; D017930:Genes, Reporter; D005822:Genetic Vectors; D008969:Molecular Sequence Data; D010957:Plasmids / Q000235:genetics*; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins; D012706:Serratia marcescens / Q000201:enzymology / Q000235:genetics*; D001616:beta-Galactosidase / Q000235:genetics
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D011993:Recombinant Fusion Proteins; D004262:DNA Restriction Enzymes; D001616:beta-Galactosidase; D001220:Aspartate Ammonia-Lyase