- Royal Veterinary and Agrcultural University, Denmark
Bacteria respond to stress by induction of specific proteins in order to be able to grow or at least to increase survival. Exposure to sublethal stress conditions therefore usually increase the tolerance of additional stress. Heat and salt stress are encountered during cheese making and the aim of this project is to investigate how these conditions affect the physiology of lactococcus lactis. The major mile stones of this project are to identify and clone the genes being induced by heat and salt, and to construct and characterize mutants in some of these genes. The effect of the stress stimulus was monitored on the protein as well as on the mRNA level. This was achieved by developing methods for labelling, extraction and 2 dimensional gel electrophoresis of lactococcal proteins. The identification of relevant protein spots is performed by western blots or by N-terminal amino acid sequencing. Also better methods for RNA extraction from lactococci have been developed. and used for isolation of heat induced genes by subtractive hybridization. Other genes were isolated by PCR techniques and the DnaK operon was a gift from M. Gasson (UK). In total the stress group at DTU and KVL now have 13 genes available, all isolated from the same lactococcal strain MG1363. Apart from the general chaperones dnaK and groELS, these genes include three different clp genes. Furthermore genes from a cell division operon have been isolated. Insertion mutants in several of these genes have been isolated and are being investigated.
|Period||01/11/93 → 28/02/97|
|Amount||2,700,000.00 Danish kroner|