Precision fermentation of milk proteins with a view to make milk and dairy products without cows


Brescó Badia, Claudia


This final thesis (hereinafter, TFG) contextualises and highlights the importance of basic laboratory research in the transformation of food strategies and resources as they are currently known; The potential of the use of bacterial cultures for the production of various proteins of great interest in the human diet, such as milk proteins, is highlighted, and this potential use is contextualised with respect to the sustainable production of these proteins without requiring the maintenance of large masses of cattle, as is currently done, with the impact that these livestock farms generate on the economy and the environment. To this end, the general objective is defined as follows: "To investigate the possibilities of efficiently inserting the milk proteins alpha-S1-casein and beta-casein by using the plasmid pEC-XC99E in the bacterial strains of Corynebacterium glutamicum wildtype ATCC 13032, M8 and JS34, taking as a reference the production and secretion of these proteins in E. coli". For this purpose, the methodological background of interest is described, establishing the theoretical bases of the importance of milk in the human diet during all age groups, the possibilities of bacterial cultures in protein production and their use during the last decades as a strategy to explore both from the approach of medicine and human nutrition, and the methodologies applied in the study, focusing specifically on homologous recombination, cloning by Gibson Assembly and the secretory expression of proteins taking advantage of the bacterial machinery and, specifically, the SEC and TAT system that transports heterologous proteins. After defining the general and specific objectives, legal aspects, ethical considerations, feasibility study and analysis of alternatives, the methodology is described, with the main theoretical foundations of the techniques applied, the materials used and the description of the protocol to be followed. Subsequently, the results obtained are described, in which, although it has been possible to transform the bacterial strains by including the plasmid pEC-XC99E, it has not been possible to obtain effective cultures in the production and secretion of any of the two proteins chosen (neither for alpha-S1-casein nor for beta-casein) in any of the three strains of C. glutamicum used in the experiments (WT, M8 and JS34). Future research strategies are discussed and presented to check whether the protein markers obtained in the histidine western blot correspond to the proteins of interest, to identify the factors that may have influenced the absence of expected results and the potential actions to achieve the general objective defined.



Ruhdal Jensen, Peter
Carnicer Heras, Marc


IQS SE - Undergraduate Program in Biotechnology