Fermentation process using marked microbial host cell strains

摘要

Presence of a contaminating microbial host cell (A), in a culture vessel containing a different host cell (B) able to use more than one carbohydrate (I), is detected by: (a) culturing (B), which includes nucleic acid encoding a particular polypeptide (II) and genetically marked so that it can not use a selected (I), (Ia), as substrate; (b) plating an isolated sample of the culture on a medium supplemented by (Ia); (c) incubating the plated cells, and (d) detecting if any colonies grow in the (Ia)-containing medium. Optionally (A) that grow in the supplemented medium may be plated on a field of different (I) to identify them from their specific (I)-utilising characteristics. The cells are yeasts or prokaryotes, particularly Gram-negative bacteria and specifically E. coli. They express an exogenous (II) and utilise e.g. maltose, ribose or rhamnose. (B) is genetically marked by making (a) a non-reversing, non-suppressing alteration in its genotype, in which case the sample is not diluted before plating or (b) a genotypic alteration that results in a carbohydrate utilisation revertant or suppressant, in which case the sample is serially diluted before plating. Optionally the strain is doubly marked by altering two (I)-utilisation pathways; (i) one type (a), in ribose utilisation and the other a reverting or suppressing mutation in rhamnose utilisation or (ii) both type (a), in maltose and ribose utilisation. Where two alterations are present, two plates are used in step (b), each supplemented with different (I). The vessel is a fermentor containing, before dilution, 108-1011 colony-forming units (cfu)/ml.