A new method for integration and stable DNA amplification in poorly transformable bacilli.
We have developed a strategy for the integration and stable amplification of DNA sequences in the chromosome of poorly transformable bacilli, which avoids the presence of a functional plasmid replication system in the integrated DNA. The parental vector for integration contains two plus origins of replication from pUB110 in the same orientation on a single plasmid. Due to the direct repeats, such plasmids produce two individual progeny vectors, one of which is dependent on the other for replication, as it lacks a functional rep gene. We have used such a progeny vector system to integrate and amplify DNA on the chromosome of Bacillus licheniformis, and show that the structure is stable in the absence of selective pressure.
Tangney
M
M
Department of Bacterial Gene Technology, Novo Nordisk A/S, Copenhagen, Denmark.
Jørgensen
P L
PL
Diderichsen
B
B
Jørgensen
S T
ST
D001407:Bacillus / Q000235:genetics*; D001412:Bacillus subtilis / Q000235:genetics; D002876:Chromosomes, Bacterial; D004269:DNA, Bacterial / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics*; D005822:Genetic Vectors; D021141:Nucleic Acid Amplification Techniques; D010957:Plasmids; D015183:Restriction Mapping; D014169:Transformation, Bacterial
D004269:DNA, Bacterial