Research On D-Latic Acid Production By Recombinant Serratia Marcescens

Lactic acid is an important organic acid, it is also an important precursor for the synthesis of a variety of substances, which is widely used in the chemical industry, agriculture, pharmaceutical industry, food industry and other fields.Requirements are lower compared with lactic acid bacteria, Serratia marcescens culture media components is relatively simple, inexpensive, has certain advantages in the conversion rate and yield indicators. The research includes culture conditions of Serratia marcescens and medium composition optimization and enhancement the expression of a key enzyme in the metabolic production of lactic acid pathway lactate dehydrogenase. As follows:Culture conditions to determine:Shake flask experiments to determine the the Serratia marcescens fermentation production of lactic acid culture conditions:temperature34℃, pH6.5,5%of the inoculum size, pre-aerobic culture accumulated the thallus late anaerobic fermentation acid production by two-stage culture methodDetermination of medium components:Through the optimization experiment, the final media components are as follows (g1-1):Yeast10;(NH)4H2PO45.0; KH2PO41.5; K2HPO4·3H2O2.0; MgSO4·7H2O1.0; FeSO4·7H2O30mg; MnSO4·7H2O5mg.Glucose content is depending on the circumstances. Optimized by the shake flask medium, lactic acid production increased from13.5±1.3g1-1to29.0±1.4g1-1.Comparison of the fermentation process:On the basis of experiments in shake flask, researching batch fermentation and fed-batch fermentation in bioreactor respectively:lactic acid production were48.5±1.6g1-1,42±2.2g1-1; lactate yields were1.35g1-1h-1,1.75g1-1h-1; the maximum cell OD600of17.8,27.4respectively; main byproduct is acetic acid and succinic acid, acetic acid.D-lactate dehydrogenase expression:Cloned lactate dehydrogenase gene, and build to express the plasmid pPbud-dldh competent cells get transferred to Serratia marcescens build over the expression strain SM R1-dldh. this strain fermentation experiments, results showed that:with the pro-strain comparison, into the plasmid, the expression amount of the lactate dehydrogenase increased to accelerate the consumption of glucose, and improves the production of lactic acid.