Breeding Of L-Lysine Producer Using Atmospheric Pressure Plasma And Its Fermentation Optimization

The high quality protein feed has required not only high content of crude protein,but also the content and composition of amino acid in feed.However,amino acids in natural feed are almost unbalanced.L-lysine,as one of the eight essential amino acids,which needs to produce feed products by exogenous addition of a certain amino acid.L-lysine,is known as the first restricted amino acid,and it not only has great application value in feed production,but also is increasing in the demand every year.At present,the production of L-lysine is mainly obtained by fermentation,and the raw materials include rice straw,wheat bran,bagasse and so on,which have the advantages of wide source and low price.Based on the atmospheric pressure room temperature plasma technology,to obtain high yield strain the L-lysine producing strains B413,B253 were mutagenic and selected.High concentration of fermentable hydrolyzed sugar was obtained by dilute acid pretreatment of corn straw,and was as carbon source of the fermentation high concentration of L-lysine was obtained by fermentation with cellulose hydrolysate.This study provided a new theoretical principle for breeding and fermentation of L-lysine strain and realizing the efficient utilization of crop straw.The main results are as follows:(1)Based on the atmospheric pressure room temperature plasma technique,the strain B413 and B253 was mutagenic and selected.The strain B413 and B253 was mutagenic for 10 times under the treatment of 100 W and 60 s.The high yield strains B413-ARTP-X-153 and B253-ARTP-X-030 were obtained respectively.The maximum yields were 12.16 g/L and 22.50 g/L,which were 10.65% and 72.94% higher than those of unmutated strains,respectively.(2)Through the study on the feeding and enzymatic hydrolysis conditions of corn straw pretreated with dilute acid,the optimum enzymatic conditions were determined as follows: water washing 2 times,feeding 4 times,enzymatic hydrolysis time 96 h,50?,150 rpm,90 g / L of glucose could be obtained at the level of shaking flask.Using this enzymatic hydrolysis condition and the enzymatic hydrolysis experiment was amplified by 30 L saccharification pot and 119.39 g / L glucose was obtained.(3)The fermentation medium was optimized by using straw hydrolysate as carbon source.The optimum combination of seed medium was obtained(g/L): glucose 30,20,Mn SO4 5,Ca CO325,Mg SO4?7H2O 0.6,K2HPO4?3H2O 1.5 g/L,Fe SO4?7H2O 4 mg/L,Mn SO4 2 mg/L and the optimum combination fermentation medium was obtained: glucose 80,corn steep liquor 15,Mn SO4 55,Ca CO3 30,Mg SO4?7H2O 0.6,K2HPO4?3H2O 1,Fe SO4?7H2O 4 mg/L,Mn SO4 2 mg/L.The optimum fermentation conditions were as follows: the first stage seed culture time was 12 h,p H 7.0,30?,180 rpm,10% to the same secondary seed.The culture medium was supplemented with 200 rpm 10% for 10 h,the p H was adjusted to 7.0 every 4 h,and the fermentation period was 84 h.The optimum medium for fermentation showed that the maximum yield of L-lysine was 28.5 g / L,which was 48.1% higher than that of the unmutated B253.