ABSTRACT
Salmonella enterica serovar Cerro 87, which was isolated from a commercial egg-producing farm, has a phosphorothioated DNA backbone resulting DNA degradation(Dnd) during the pulsed field gel electrophoresis (PFGE) process. In this research, a gene deletion mutant XTG103 was engineered with the entire dnd gene cluster knocked out by double crossover using vector pKOV-kan, and lost Dnd phenotype corre- spondingly. We regulated the DNA phosphorothioation by heterogenous expression of dnd gene cluster with an isopropyl ?-D-1-thiogalactopyranoside (IPTG) inducible promoter PlacZ.
ABSTRACT
pHZ1080, an E. coli-Streptomyces shuttle expression vector was constructed in order to explore the utilization of lambda phage regulated expression elements in Streptomyces. A 2.7 kb polyketide synthase (PKS) gene from Streptomyces sp. FR-008 was inserted into downstream of lambda phage promoter (PR) to give the shuttle plasmid, pHZ1067. The PKS protein was expressed in Streptomyces lividans carrying pHZ1067 in a heat-dependent manner, as it did in E. coli. The PKS protein expressed in both hosts with same molecular weight was detected by SDS-PAGE and Western-blot. The successful heat-induced expression of PKS suggested that pHZ1080 was useful and convenient for heat-induced expression of heterologous genes in both E. coli and Streptomyces.