Cloning of prochymosin gene in lactococcus lactis bacteria

In recent years the increase in milk production worldwide on one hand and reduction in natural sources of "rennin" on the other, has resulted in significant decrease in rennin-produced cheese. In this research, it was attempted to clone the caprin prochymosin gene in the lactococcus lactis. If succeeded, the results of this work would allow production of cheese without the need for adding exogenous rennin or low quality fungal enzymes. The prochymosin gene which had been previously cloned in pET plasmid in E. coli BL21 was amplified by PCR and was then isolated from the agar gel, ligated to plasmid PTZ and cloned in E. coli GM. The prochymosin gene was cut by enzyme and ligated to L. lactis plasmid PNG and cloned in E. coli MC1061. The plasmid then isolated from E. coli MC1061 and transferred to L. lactis NZ9000 and recombinant cloned were sequenced.