STABLE HIGH-COPY-NUMBER BACTERIOPHAGE-LAMBDA PROMOTER VECTORS FOR OVERPRODUCTION OF PROTEINS IN ESCHERICHIA-COLI

The construction of new high-copy-number (hcn) lambda-promoter express ion vectors is described. All these vectors (1) contain tandem lambda p(R) and p(L) promoters upstream of an extensive multiple cloning site (MCS) for insertion of genes, (2) direct expression of the lambda cI( ts)857 gene, enabling their use in any Escherichia coli host strain fo r thermal induction of gene overexpression, and (3) bear the par locus of plasmid pSC101, ensuring their stable maintenance at hen in the ab sence of continuous antibiotic selection. Six of the vectors also cont ain efficient ribosome-binding sites upstream of unique HpaI or NdeI s ites in their MCS regions, and two contain sequences that encode N-ter minal poly-His. The performance of these vectors was assessed by using them to overproduce the E. coli HMP flavohaemoprotein and the bacteri ophage M13 gene II replicator protein.