Abstract: The invention relates to modifying plasmid origins of replication to create hybrid origins of replication containing nucleotide sequences from more than one plasmid. The invention also relates to a modified origin of replication cassette that is portable or exchangeable due to the creation of multiple cloning sites flanking the origin of replication. Methods and plasmids for use in exchanging origins of replication are disclosed. Such modified or hybrid plasmids provide useful cloning tools that allow for regulation of the level of expression of a desired protein.

Abstract: The instant invention is drawn to the methods and compositions necessary to provide recombinant proteins with a substantially reduced or eliminated content of norleucine or other non-standard amino acids. Various embodiments of the invention provide for the substantial elimination of the incorporation of non-standard amino acids into recombinant proteins by the co-expression or enhanced expression of a protein (or the enzymatically active portion thereof) capable of degrading norleucine or other non-standard amino acids, including norvaline, beta-methylnorleucine, and homoisoleucine. In certain particular embodiments of the invention, the norleucine is degraded by a glutamate dehydrogenase, a leucine dehydrogenase, a valine dehydrogenase, a phenylalanine dehydrogenase, a glutamate/leucine/phenylalanine/valine dehydrogenase, or an opine dehydrogenase. Also provided are the cells and DNA constructs for carrying out these methods.

Abstract: The invention relates to modifying plasmid origins of replication to create hybrid origins of replication containing nucleotide sequences from more than one plasmid. The invention also relates to a modified origin of replication cassette that is portable or exchangeable due to the creation of multiple cloning sites flanking the origin of replication. Methods and plasmids for use in exchanging origins of replication are disclosed. Such modified or hybrid plasmids provide useful cloning tools that allow for regulation of the level of expression of a desired protein.

Abstract: The invention allows improved expression of heterologous polypeptides such as bovine somatotropin (bST). Novel compositions and methods are provided for production of bST from a native bST cDNA in transformed host cells such as E. coli. In particular, DNA segments comprising novel ribosome binding sites and adjacent sequences are provided, which in conjunction with a promoter sequence direct high level expression of bST from its native cDNA. The invention also provides expression constructs comprising the ribosome binding sites and methods for the use of such constructs.