Development of a method for heterologous gene expression in Enterobacter amnigenus, a potential host for the biological control of mosquito larvi.
An integrative plasmid containing a 1.3 kb fragment of chromosomal DNA from Enterobacter amnigenus was constructed. The Omega fragment encoding spectinomycin/streptomycin resistance was cloned into the unique BglII site of the resulting plasmid, and the interrupted fragment was transferred via plasmid pMAK705 by electroporation into E. amnigenus with a selection for spectinomycin resistance. Cointegrants were resolved to generate an E. amnigenus strain that expressed spectinomycin resistance, but grew as rapidly as the parental strain. The cloned fragment encodes a putative homologue of the proW gene of Escherichia coli that is not essential for E. amnigenus growth. The integrative plasmid is now available to introduce any heterologous DNA into the E. amnigenus chromosome, for the construction of promoter-probe vectors for the studies of gene regulation, or to construct plasmids suitable for the isolation of secretion signals. Immediate applications of this system will include the expression and secretion of crystal toxins from bacilli for the biological control of mosquito larvae infected with the bacterial host.
Nantapong
Nawarat
N
Institute of Molecular Biology and Genetics, Mahidol University, Salaya Campus, Nakornpathom, 73170 Thailand.
Tanapongpipat
Sutipa
S
Cole
Jeffrey
J
Panyim
Sakol
S
D018528:ATP-Binding Cassette Transporters / Q000235:genetics; D000900:Anti-Bacterial Agents / Q000096:biosynthesis; D001426:Bacterial Proteins / Q000235:genetics; D003001:Cloning, Molecular / Q000379:methods*; D004351:Drug Resistance / Q000235:genetics; D004754:Enterobacter / Q000187:drug effects / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics; D029968:Escherichia coli Proteins; D015870:Gene Expression; D005822:Genetic Vectors; D008565:Membrane Proteins / Q000235:genetics; D008969:Molecular Sequence Data; D009032:Mosquito Control; D010957:Plasmids; D000198:Spectinomycin / Q000096:biosynthesis; D013307:Streptomycin / Q000096:biosynthesis
D018528:ATP-Binding Cassette Transporters; D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D008565:Membrane Proteins; C490210:ProW protein, E coli; D000198:Spectinomycin; D013307:Streptomycin