| Microbial fertilizers can effectively improve soil fertility,enhance plant disease resistance,improve crop quality,accord with the production concept of green environmental protection,and have broad prospects for development.The integrated technology of water and fertilizer has the advantages of fast fertilizer efficiency,high nutrient utilization rate,water and fertilizer saving,reducing production cost and benefiting environmental protection.It is an important technology in modern agriculture.It is of great significance to study the combination of microbial fertilizer and water and fertilizer integration technology.Bacillus subtilis HG-15 was isolated from wheat rhizosphere soil in saline-alkali land of the Yellow River delta by this laboratory.This strain has broad-spectrum antibacterial activity,can produce auxin(IAA)and ACC deaminase,and has the dual function of disease prevention and growth promotion.It can colonize the rhizosphere of wheat,corn,cucumber and poplar stably and has good salt tolerance.As a microbial fertilizer strain,this strain has been widely used in production.In order to study the application of this strain in the integration of water and fertilizer,the fully soluble medium formula and fermentation conditions for the fermentation of this strain were optimized in this study.The main findings are as follows:1.The basic medium of Bacillus subtilis HG-15 was screened by shaking flask fermentation,and the optimal basic medium formula was obtained:Glucose 4.5g/L,Yeast powder 5.0g/L,peptone 10.0g/L,NaCl 10.0g/L,MgSO4 0.2g/L,MnSO4 0.2g/L,water 1.0L.The number of bacteria in the fermentation broth is 2.076×10~9 cfu/mL.2.Through single factor experiment and orthogonal experiment,the fermentation medium formulation of HG-15 strain was preliminarily optimized.The single factor test determined that the best carbon source was glucose,the best nitrogen source was yeast powder,and(NH4)2SO4;the best carbon-nitrogen ratio and the best inorganic salt combination were determined by orthogonal test.The medium formula obtained through preliminary optimization is as follows:Glucose 4.5g/L,Yeast powder 46.5g/L,(NH4)2SO40.4g/L,NaCl 10.0g/L,MgSO4 0.8g/L,MnSO4 0.2g/L,KH2PO4 0.3 g/L,water 1.0L.Shake flask fermentation with this formula,the number of bacteria in the fermentation broth is2.93×10~9 cfu/mL.3.Through the single factor test,the optimal conditions for the fermentation of the HG-15 strain were:inoculation volume of 10%,filling volume of 50mL/250mL,temperature of 28?,rotation speed of 200r/min.Under this condition,the initial optimized medium formula was used for shake flask fermentation,and the number of bacteria in the fermentation broth is 4.167×10~9cfu/mL.4.Under the optimal fermentation conditions,the response surface methodology optimization experiment was carried out on the fermentation medium formulation of HG-15strain.The results of the Plackett-Burman test showed that the significant factors affecting the number of bacterial in the fermentation broth as yeast powder,NaCl and MnSO4;the optimal concentration of significant influencing factors was determined by the Steepest ascent design and Central composite design,and the optimal fully soluble medium formula of HG-15 strain was finally obtained as follows:Glucose 4.5g/L,Yeast powder 60.125g/L,(NH4)2SO4 0.4g/L,NaCl 9.0g/L,MgSO4 0.8g/L,MnSO4 0.35g/L,KH2PO4 0.3 g/L,water 1.0L.Use the optimized fully soluble medium formulation of the HG-15 strain fermentation obtained by optimization,and shake the flask under the optimal fermentation conditions.When the spore rate in the fermentation broth was more than 90%,the number of bacteria is5.122×10~9cfu/mL,compared with the basic medium,the number of bacteria in the fermentation broth was increased by 2.47 times.The fully soluble medium formula and fermentation conditions of Bacillus subtilis HG-15 obtained in this study have higher reference value for the industrial fermentation of this strain and its application in the integration of water and fertilizer. |
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