RRC ID 33455
Author Simons RW, Houman F, Kleckner N.
Title Improved single and multicopy lac-based cloning vectors for protein and operon fusions.
Journal Gene
Abstract We describe several new vectors for the construction of operon and protein fusions to the Escherichia coli lacZ gene. In vitro constructions utilize multicopy plasmids containing suitable cloning sites located between upstream transcription terminators and downstream lac operon segments whose lacZ genes retain or lack translational start signals. Single-copy lambda prophage versions of multicopy constructs can be made genetically, without in vitro manipulation. The new vectors, both single and multicopy, are improved in that they have very low levels of background lac gene expression, which makes possible the easy detection and accurate quantitation of very weak transcriptional and translational signals. These vectors were developed for analysis of the expression of IS10's transposase gene, which is transcribed less than, once per generation, and whose transcripts are translated on average less than once each. Both single and multicopy constructs can also be used to select mutations affecting fusion expression, and mutations isolated in single-copy constructs can be crossed genetically back onto multicopy plasmids for further analysis.
Volume 53(1)
Pages 85-96
Published 1987-1-1
DOI 10.1016/0378-1119(87)90095-3
PII 0378-1119(87)90095-3
PMID 3596251
MeSH Gene Expression Regulation Genetic Vectors* Lac Operon* Operon Protein Biosynthesis Recombinant Fusion Proteins / genetics* Recombinant Proteins / genetics* Transcription, Genetic
IF 2.984
Times Cited 1465
Prokaryotes E. coli pRS414 pRS415 pRS551 pRS552