Gist-brocades N.v.

Dsm N.v.

Gist-brocades B.v.

Leo J Mulleners

Peter Michael Andreoli

Johan Pieter Sanders

Yvonne J Vos

Johannes C Van Der Laan

Jan H Van Ee

Christiaan A Van Eekelen

Johannes Abel Van Den Berg

Johannes A Van Den Berg

Transformed prokaryotic hosts are provided comprising two or more copies of a DNA sequence stably maintained in their chromosome, said DNA sequence comprising a gene encoding a polypeptide of interest wherein said copies are separated by endogenous chromosomal DNA sequences. Methods are also provided for producing said transformed host strains. The transformed host strains are capable of increased production of the polypeptide of interest compared to host strains which already produce said polypeptide. Preferred host strains are Bacillus novo species PB92 which produces a high-alkaline proteolytic enzyme and B. licheniformis strain T5 which produces a thermostable .alpha.-amylase, and mutants and variants of said strains. Preferred poly-peptide encoding genes are the protease encoding gene obtainable from Bacillus PB92 and the .alpha.-amylase encoding gene obtainable from B. licheniformis strain T5.

Stable gene amplification in chromosomal DNA of prokaryotic

Novel methods and novel industrial unicellular microorganism strains, particularly industrial Bacillus strains, are provided for enhanced production of endogenous and exogenous polypeptides. Cloning vehicles containing the gene expressing the polypeptide of interest are introduced into a compatible host. Transformed hosts harboring the introduced vehicle in a stable way by integration of the vehicle into the host cells chromosome are selected. Efficient transfer of the vehicle containing the gene of interest is achieved, with the resulting industrial strain transformants being effective, stable producers of the desired polypeptide product.

Transformed industrial bacillus strains and methods for making and using

Transformed prokaryotic hosts are provided comprising two or more copies of a DNA sequence stably maintained in their chromosome, said DNA sequence comprising a gene encoding a polypeptide of interest wherein said copies are separated by endogenous chromosomal DNA sequences. Methods are also provided for producing said transformed host strains. The transformed host strains are capable of increased production of the polypeptide of interest compared to host strains which already produce said polypeptide. Preferred host strains are Bacillus novo species PB92 which produces a high-alkaline proteolytic enzyme and B. licheniformis strain T5 which produces a thermostable .alpha.-amylase, and mutants and variants of said strains. Preferred polypeptide encoding genes are the protease encoding gene obtainable from Bacillus PB92 and the .alpha.-amylase encoding gene obtainable from B. licheniformis strain T5.

Novel methods and novel industrial unicellular microorganism strains, particularly industrial Bacillus strains, are provided for enhanced production of endogenous and exogenous polypeptides. Cloning vehicles containing the gene expressing the polypeptide of interest are introduced into a compatible host. Transformed hosts harboring the introduced vehicle in a stable way by integration of the vehicle into the host cells chromosome are selected. Efficient transfer of the vehicle containing the gene of interest is achieved with the resulting industrial strain transformants being effective, stable producers of the desired polypeptide product.

Novel methods and novel industrial unicellular microorganism strains, particularly industrial Bacillus strains, are provided for enhanced production of endogenous and exogenous polypeptides. Cloning vehicles containing the gene expressing the polypeptide of interest are introduced into a compatible host. Transformed hosts harboring the introduced vehicle in a stable way by integration of the vehicle into the host cells chromosome are selected Efficient transfer of the vehicle containing the gene of interest is achieved, with the resulting industrial strain transformants being effective, stable producers of the desired polypeptide product.

Molecular cloning and expression in industrial Bacillus species

Novel methods and novel industrial Bacillus strains are provided for enhanced production of serine protease. Plasmid constructs comprising the serine protease gene are introduced into a compatible host, generally a Bacillus already overproducing serine protease. Preferred host cells are those from Bacillus novo sp. PB92 and the preferred serine protease gene also originates from Bacillus PB92. Integration and maintenance of the plasmid in the host cell chromosome can be achieved by including a temperature sensitive origin of replication in the plasmid construct and growing under selective conditions.

Growing community of inventors

Rijen, Netherlands

Leo J Mulleners