Dh5α t1r e coli cells

Example 6

The starting plasmids used for the cloning and expression of the cyclodextrin glycosyltransferase (CGTase) gene from Klebsiella pneumoniae M5a1 (Genebank No. M15264) were again the plasmid pMT1 and the plasmid pCGT described in US2008076158 A1.

For the generation of a new production plasmid for the production of CGTase, based on pMT1, a MauBI-BsaI fragment from the plasmid pCGT, which codes for the CGTase gene from Klebsiella pneumoniae M5a1, was ligated with a 4004 bp MauBI-BsaI fragment from the plasmid pMT1. Said 4004 bp fragment from the plasmid pMT1 codes for the ColE1 ori, the lac/tac operator and the tetracyline resistance gene (tetR).

The ligation preparation was transformed into “DH5α™-T1R E. coli cells” (Life Technologies GmbH), multiplied in said cells, and the DNA sequence of the isolated plasmids was verified by means of sequencing. The resulting expression plasmid has the designation pCGT_tetR (see FIG. 6).

Example 7

The starting plasmids used for the cloning and expression of the genes for the anti-lysozyme Fab fragment were again the plasmid pMT1 and the pFab-anti-lysozyme described in US20080076158 A1.

For the generation of a new production plasmid for the production of the antibody fragment Fab-anti-lysozyme, based on pMT1, a MauBI-BsaI fragment from the plasmid Fab-anti-lysozyme, which codes for the two chains, i.e., the heavy chain (V_H-C_H1 domains) and the light chain (V_L-C_L domains) of the anti-lysozyme Fab fragment, was ligated with a 4004 bp MauBI-BsaI fragment from the plasmid pMT1. Said 4004 bp fragment from the plasmid pMT1 codes for the ColE1 ori, the lac/tac operator and the tetracyline resistance gene (tetR).

The ligation preparation was transformed into “DH5α™-T1R E. coli cells” (Life Technologies GmbH), multiplied in said cells, and the DNA sequence of the isolated plasmids was verified by means of sequencing. The resulting expression plasmid has the designation pFab-anti-Lysozyme_tetR (see FIG. 7).