Study On The Method And System Design Of Mutagenesis For Probiotics By Atmospheric And Room Temperature Plasma

With the improvement of living standards,the demand for microorganisms is also rising,but the production performance of many existing strains can no longer meet the production demand.In order to obtain more excellent strains,the existing strains must be improved.At present,the improvement of strains is mainly obtained by mutation.In order to achieve efficient and convenient mutagenesis,this paper takes Bacillus velezensis and Lactobacillus as examples,uses low-temperature plasma produced by high-voltage discharge technology to mutagenize them.Then a set of mutagenic system was designed which use the plasma discharge in gas bubble.(1)A high voltage corona electric field to produce plasma was used to mutate Bacillus velezensis,the mechanism of mutation was investigated and the optimal mutation conditions were obtained.According to the results,after treatment with different high voltage electric fields and treatment time,the total number of colonies and survival rate of Bacillus velezensis have changed greatly,and the high voltage electric fields have positive and negative effects on the growth of Bacillus velezensis.At the same time,the survival state of bacteria also has a great impact on its mutagenic effect.The mutagenic effect of plasma on a single colony is the best,the mutagenic effect on bacterial liquid is the second,and the mutagenic effect on bacterial powder is the worst.After mutagenesis,it was observed that the morphology and number of colonies changed after high-voltage electric treatment.The higher the voltage,the more irregular the morphology and the less the number of colonies.With the increase of time,the morphology and number of colonies increased first and then decreased.(2)Plasma treatment will have some effect on the enzyme production performance of Bacillus.Different high-voltage electric field treatments have different effects on the amylase activity in the bacterial solution,and the single colony enzyme activity of the same high-voltage electric field treatment is also significantly different.After partial treatment,the amylase activity of mutant strains increased significantly.Under the optimum input energy,the corona discharge treatment can improve the activity of Bacillus belles.The sequence alignment analysis of 16 S r RNA also confirmed that the corona discharge treatment caused the mutation of Bacillus belles.Compared with the original strain,the enzyme activity of mutant bacteria is greatly improved,which is a positive effect and can meet the production requirements.(3)In order to obtain higher mutagenic efficiency,a mutagenic system using water bubble discharge to produce plasma was designed after various factors affecting the complex building,and the system was verified with Lactobacillus plantarum.The experiment showed that the new strain with high antibacterial activity and protease activity was screened out through the mutagenic system of water bubble discharge to produce plasma.After mutagenic culture,the lactic acid bacteria colony is larger than the control group,the surface of the colony is drier,and the edge of the colony becomes irregular.Although the number of live bacteria of Lactobacillus plantarum obtained by mutation is small,its OD value is higher than that of the control group,its p H value is lower than that of the control group,the diameter of the bacteriostatic ring is larger,and the activity of protease is higher,which indicates that its performance is better.It can be considered that it grows faster and dies faster.In order to verify its genetic stability,it was subcultured.According to the results of subculture,it can be determined that they can be stably inherited.(4)According to the test data,this mutation system uses DC high voltage mutation,the mutation process is simple to operate,and the mutation efficiency is high.Bacterial mutagenesis is carried out by means of bubble discharge in liquid,which greatly increases the dose of the mutated sample receiving the action of plasma and increases the probability of being mutated.At the same time,due to the existence of bubbles,the bacterial solution is disturbed,which makes up for the difference with other physical mutagenic methods(such as α X ray,ultraviolet ray,particle beam,etc.).Compared with other existing plasma mutagenic equipment,this mutagenic system is characterized by small size,light weight,low cost,low energy consumption,simple operation,high mutagenic efficiency and convenient transportation.