Study On The Funcion Of Lactobacillus Plantarum C3005IN Reducing Cholesterol And Construction Of The Mutation

Cerebrovascular disease is the number one cause of death to the people. High cholesterol content in people is considered to be the main hazardous factors evoked cardiovascular and cerebrovascular diseases. Therefore, study on cholesterol-lowering become more popular in recent years. The probiotic lactic acid bacteria, deriving from security and a wealth of sources, are widely accepted for people.In recent years the phage Mμ DN A transposition recombination reaction is a rise of new technology to study bacterial functional genomics. It’s used by many scholars because of the characteristics of a single point of randomly inserted.We used Lactobacillus plantarum C3005as the research object in this experiment.Firstly, we study on its cholesterol-lowering function. The results showed that Lactobacillus plantarum C3005cell growth was positively related to cholesterol removal rate. In the4to12hours, since the bacteria in the logarithmic phase, the cholesterol removal rate increased. When the cholesterol concentration of medium was30mg/100mL, the cholesterol removal rate of bacterial reach to maximum,60.3%. In the experiment, both of the resting cells and dead cells could reduce the content of cholesterol in the culture, and different concentrations of bacterial cells also had a significant effect on their cholesterol removal when bile salt was added in medium, the cholesterol removal rate of the bacterial cells decreased.Secondly, the electroporation conditions of Lactobacillus plantarum C3005were optimized, and the highest transformation efficiency was obtained. The shuttle plasmid pMG36e extracted from E.coli was transformed to Lactobacillus plantarum C3005by electroporation. The conversion efficiency was studied, and the experiments were conducted on the conditions of the period of cell growth, the growth medium components, the storage temperature of the bacterial recovery medium components, recovery training time and plasmid concentration. It indicated that the highest transformation efficiency,1.19×105transformants μg-1of input plasmid DNA could be achieved when cells were grown to an OD600of0.5, the final concentration of sucrose in recovery medium was added to0.1mol/L, the training time for recovery was2.5hours. In addition,0.7M NaCl was added in MRS medium for pretreatment, when the concentration of the plasmid was0.5ng/μL, Lg (transformants/μg plasmid DNA) was6.14. Lay the foundation for the high transformation efficiency of transposition complexes.Finally, based on the phage Mμ DNA transposition reaction, we built the Lactobacillus plantarum C3005mutants library. Amplification, purification and concentrated the transposable DNA to make the concentration became higher in order to achieve the required value for the experiment. Then prepared transposition complexes and used the optimized electroporation conditions to transformed.72transformants were obtained.