Airlift fermentation method of bacillus thuringiensis

Abstract

The invention discloses an airlift fermentation method of bacillus thuringiensis, which is free of stirring in the fermentation process and comprises the following steps: S1, taking a bacillus thuringiensis seed solution and a fermentation culture medium with the inoculation amount of 0.5-10 wt%; s2, adding the seed solution and the fermentation culture medium into an airlift fermentation tank, controlling the charging coefficient to be 60-80%, the ventilatory capacity to be 0.6-0.8 vvm and the defoaming agent to automatically control defoaming, and carrying out fermentation culture for 50-70 hours at the pH of 6.5-7.5 and the temperature of 28-31 DEG C; and S3, putting into a tank to obtain the bacillus thuringiensis fermentation liquor. In the fermentation process, the fermentation liquor with the same or better effect as the fermentation liquor under stirring can be obtained without stirring.

Description

Technical Field

[0001] The invention relates to the field of microbial fermentation, and in particular to an airlift fermentation method of Bacillus thuringiensis. Background Art

[0002] In the agricultural field, some strains of Bacillus thuringiensis (Bt) can effectively deal with some pests or diseases, such as aphids and powdery mildew, by producing specific antibiotics or enzymes. The advantages of this microbial insecticide are that it is environmentally friendly, harmless to humans and beneficial insects, degradable, and will not accumulate in the environment, ultimately forming a green and harmless agricultural control method. In addition, due to these characteristics of microbial insecticides, they are of great value in preventing pests from developing resistance to chemical insecticides. Diversified agricultural control methods can better maintain ecological balance and protect the growth of crops.

[0003] Bacillus thuringiensis has very little impact on humans and non-target organisms because the intestinal environment (acidic environment) of humans and non-target organisms cannot activate crystal proteins, and there are no receptors on their intestinal epithelial cells that the toxin can bind to, so they will not be affected by the toxin. This makes Bacillus thuringiensis an ideal biopesticide.

[0004] Bacillus thuringiensis kurstaki was introduced from Russia and has a wide antibacterial spectrum and high inhibition rate. Its fermentation liquid and the preparations prepared by using it are used to control lepidopteran pests and have good effects.

[0005] In the fermentation broth of Bacillus thuringiensis, the amount of spores and protein content are related to the effect of controlling lepidopteran pests. The higher the amount of spores and protein content, the better the effect.

[0006] In the existing Bacillus thuringiensis fermentation process, some manufacturers transfer the Bacillus thuringiensis seed liquid into a fermentation tank with stirring for fermentation and culture, and obtain the fermentation liquid after the tank is put into use.

[0007] The stirred fermentation of Bacillus thuringiensis has the following defects: high energy consumption, energy consumption (air oxygen supply, steam sterilization, stirring power consumption, circulating water cooling) accounts for 20% to 30% of the total product cost, of which stirring accounts for 20% to 40% of the total energy consumption.

[0008] Therefore, how to reduce energy consumption without reducing the effect of fermentation liquid has become an urgent problem to be solved in the industry.

[0009] The above background technology is for facilitating the understanding of the present invention and is not a known technology disclosed to the general public before the application of the present invention. Summary of the invention

[0010] In view of the above-mentioned defects, the present invention provides a Bacillus thuringiensis airlift fermentation method, which can obtain a fermentation liquid with the same or better effect as that under stirring without stirring during the fermentation process.

[0011] The technical solution is: a Bacillus thuringiensis airlift fermentation method, without stirring during the fermentation process, comprising the following steps:

[0012] S1, taking Bacillus thuringiensis seed solution and fermentation medium, the inoculation amount is 0.5-10wt%;

[0013] S2, adding seed liquid and fermentation medium into an airlift fermenter, with a loading factor of 60-80%, aeration of 0.6-0.8 vvm, automatic defoaming control by defoaming agent, pH 6.5-7.5, and fermentation culture at 28-31°C for 50-70 hours;

[0014] S3, the Bacillus thuringiensis fermentation liquid is obtained in the lower tank.

[0015] Furthermore, in S2, when the fermentation culture reaches pH rebound, 30-60 wt % glucose solution is continuously added to control the dissolved oxygen at 20-30%.

[0016] Furthermore, in S2, 50 wt% glucose solution is added until 5 to 15 hours from the end of the fermentation, and then the addition of 30 to 60 wt% glucose solution is stopped.

[0017] Furthermore, the fermentation medium comprises:

[0018] Yeast powder 1-10wt%, glucose 1-5wt%, potassium dihydrogen phosphate 0.1-0.5wt%, magnesium sulfate 0.08-0.2wt%, calcium carbonate 0.1-0.5wt%, sodium chloride 0.01-0.05wt%, dapoxetine 0.02-0.08wt%, and the balance is water.

[0019] Furthermore, the fermentation medium comprises:

[0020] 1-10wt% soy protein powder, 1-5wt% molasses, 0.1-0.5wt% potassium dihydrogen phosphate, 0.08-0.2wt% magnesium sulfate, 0.1-0.5wt% calcium carbonate, 0.01-0.05wt% sodium chloride, 0.02-0.08wt% folate, and the balance is water.

[0021] Further, in S1, the Bacillus thuringiensis seed liquid is mainly prepared by the following steps:

[0022] S11, Bacillus thuringiensis kurstaki was activated on plates containing LB medium and cultured at 28-31°C for 12-36 h;

[0023] S12, picking a single colony and inoculating it into a shake flask filled with shake flask culture medium, and culturing it in a shaker at 28-31°C and 100-500 r / min for 12-36 hours to form a seed solution;

[0024] S13, the seed liquid is inoculated into a seed tank filled with a seed tank culture medium, and cultured at 28-31°C and 100-500r / min for 12-36h to form a pre-fermentation seed liquid, with a charging factor of 60-80%, an actual digestion time of 20-50min, an inoculation amount of 0.5-10wt%, and an aeration volume of 0.6-0.7vvm.

[0025] Furthermore, in S11, the components of the LB culture medium are: 5-15g of tryptone, 2-8g of yeast extract, 5-15g of sodium chloride, and 10-20g of agar powder per 1000ml of distilled water; in S12, the components of the shake flask culture medium are: 0.5-1.5wt% of tryptone, 0.2-0.8wt% of yeast extract, 0.5-1.5wt% of sodium chloride, and the remainder is water; in S13, the components of the seed culture medium are: 1-5wt% of yeast powder, 1-3wt% of glucose, 0.1-0.3wt% of potassium dihydrogen phosphate, 0.1-0.2wt% of magnesium sulfate, 0.2-0.4wt% of calcium carbonate, 0.02-0.04wt% of sodium chloride, 0.02-0.08wt% of folate, and the remainder is water.

[0026] Furthermore, the Bacillus thuringiensis is Bacillus thuringiensis kurstaki.

[0027] Furthermore, the airlift fermentation tank includes a tank body, which is provided with a feed inlet, a discharge port, and a feed and air passage port, the tank body forms an inner cavity of the tank body, the feed inlet is opened at the top of the tank body, the discharge port is opened at the bottom of the tank body, the feed and air passage ports are located at the lower part of the tank body, the feed and air passage ports are connected to a main pipe, the main pipe passes through the tank body, a part of the main pipe is located in the inner cavity of the tank body, and the remaining part is located outside the inner cavity of the tank body, the end of the main pipe located outside the inner cavity of the tank body is connected with an air inlet pipe and a feed pipe, a feed valve and a feed cup are installed on the feed pipe, an air inlet valve is installed on the air inlet pipe, an inner cylinder is installed in the inner cavity of the tank body, the top of the inner cylinder is provided with an upper opening, the bottom is provided with a lower opening, the inner cylinder forms an inner cavity of the inner cylinder, the main pipe is located Part of the inner cavity of the tank body passes through the inner cylinder and is located in the inner cavity of the inner cylinder. Its end opening is downward and is connected to the distributor. A first pushing unit is arranged below the bottom of the distributor. More than three horizontal distribution pipes are connected to the side wall of the distributor. The more than three horizontal distribution pipes are evenly distributed along the circumference. More than two second pushing units are arranged above each horizontal distribution pipe. The first pushing unit is located below the bottom of the inner cylinder, and the second pushing unit is located in the inner cavity of the inner cylinder. The first pushing unit is connected to the distributor through a connecting pipe, and the second pushing unit is connected to the horizontal distribution pipe through a connecting pipe. The pushing inlet of the first pushing unit is at the top and the pushing outlet is at the bottom, and the pushing inlet of the second pushing unit is at the bottom and the pushing outlet is at the top.

[0028] Compared with the prior art, the present invention has the following beneficial effects:

[0029] In order to save energy, the present invention adopts a gas lift tank for the fermentation process. On this basis, in order to obtain the same fermentation effect as that of a stirred fermenter, the fermentation process parameters and process conditions are changed, and finally the fermentation effect is achieved without stirring, that is, the same fermentation effect as that of a stirred fermenter or even better effect. DETAILED DESCRIPTION

[0030] The present invention will be further described below.

[0031] Figure 1 Schematic diagram of the structure of the fermentation tank used in the present invention, the fermentation tank comprises a tank body 1, a feed port 2, a discharge port 3 and a feed and air passage port 4 are provided on the tank body 1, the tank body 1 forms a tank inner cavity 7, the feed port 2 is provided at the top of the tank body 1, the discharge port 3 is provided at the bottom of the tank body 1, the feed and air passage port 4 is located at the lower part of the tank body 1, the feed and air passage port 4 is connected with a main pipe 5, the main pipe 5 passes through the tank body 1, a part of the main pipe 5 is located in the tank inner cavity 7, and the rest of the main pipe 5 is located outside the tank inner cavity 7, the end of the main pipe 5 located outside the tank inner cavity 7 is connected with an air inlet pipe 51 and a feed pipe 52, a feed valve 53 and a feed cup 54 are installed on the feed pipe 52, an air inlet valve 55 is installed on the air inlet pipe 51, an air compressor (not shown in the figure) is connected to the air inlet end of the air inlet pipe 51, a cooling coil (not shown in the figure) and an inner tube 6 are installed in the tank inner cavity 7, and an upper opening 61 is provided on the top of the inner tube 6 , a lower opening 62 is opened at the bottom, the inner tube 6 forms an inner tube cavity 8, the part of the main pipe 5 located in the inner cavity 7 of the tank body passes through the inner tube 6, is located in the inner tube cavity 8, and its end opening is downward and is connected to the distributor 9. A first pushing unit 10 is arranged below the bottom of the distributor 9, and more than three horizontal distribution pipes 11 are connected to the side wall of the distributor 9. The more than three horizontal distribution pipes 11 are evenly distributed along the circumference, and more than two second pushing units 12 are arranged above each horizontal distribution pipe 11. The first pushing unit 10 is located below the bottom of the inner tube 6, and the second pushing unit 12 is located in the inner tube cavity 8. The first pushing unit 10 is connected to the distributor 9 through the connecting pipe 13, and the second pushing unit 12 is connected to the horizontal distribution pipe 11 through the connecting pipe 13. The pushing inlet of the first pushing unit 10 is at the top, and the pushing outlet is at the bottom, and the pushing inlet of the second pushing unit 12 is at the bottom, and the pushing outlet is at the top.

[0032] The feed valve 53 and the air inlet valve 55 are both electromagnetic valves. The cooling coil and the tank body 1 are the same as those in the prior art. The positions of the feed port 2 and the discharge port 3 are the same as those in the prior art. In the prior art, the cooling coil takes away the heat generated in the reaction to maintain the reaction temperature. The main pipe 5 passes through the feed and air through port 4, and the main pipe 5 and the tank body 1 are fixed by welding at the feed and air through port 4. The bottom of the inner tube 6 is fixed by a support, and the bottom of the support is connected to the inner wall of the bottom of the tank body 1.

[0033] Furthermore, there are eight horizontal distribution pipes 11 , and two second pushing units 12 are installed on each distribution pipe 11 .

[0034] Furthermore, the main bodies of the first pushing unit 10 and the second pushing unit 12 are both cylinders, one end of the cylinder is connected to the pushing inlet, and the other end is connected to the pushing outlet.

[0035] Furthermore, the push inlet and the push outlet are both conical structures, the small ends of the push inlet and the push outlet are connected to the cylinder, and the large ends are far away from the cylinder.

[0036] Furthermore, the diameter of the portion of the connecting pipe 13 located in the push inlet gradually decreases along the gas running direction.

[0037] Furthermore, the pushing outlet of the first pushing unit 10 is adjacent to the discharge port 3 and is located above the discharge port 3 .

[0038] Furthermore, the upper opening 61 is in the shape of an inverted truncated cone.

[0039] Before production, production materials enter the tank body cavity 7 and the inner tube cavity 8 through the feed port 3. During production, compressed gas enters from the air inlet pipe 51 through the main pipe 5, and runs through the first pushing unit 10 and the second pushing unit 12. Among them, the second pushing unit 12 diffuses and flows upward, and circulates from the inside to the outside in a directional manner, forming an inside-up and outside-down. The first pushing unit 10 disturbs and mixes the material retained at the bottom of the tank body cavity 7, and the rising gas drives the material in the retention area to the inner tube cavity 8.

[0040] In the present invention, the protein content detection method is as follows: take 2 ml of fermentation liquid and centrifuge at 10000 rpm for 10 min, wash the precipitate twice with sterile water, wash the precipitate twice with 0.5 mol / L NaCl; add 1 ml of lysis solution (50 mM Na2CO3, 10 Mm disodium EDTA, add mercaptoethanol with a final concentration of 3% before use, and adjust the pH to 9.5) to the precipitate, and lyse at 4°C for 5-6 hours; centrifuge at 10000 rpm for 10 minutes, take the supernatant and add 2 ml of 4 M NaAc-Hac (pH = 4.5), and precipitate at 4°C for 4 hours. Centrifuge at 4°C and 10000 rpm for 20 minutes, wash the precipitate with sterile water, and then dissolve it in 2 ml of 50 mM Na2CO3 solution with a pH of 9.5, and measure the protein content using an ultra-micro nucleic acid protein detector.

[0041] Example 1 Preparation of Seed Liquid before Fermentation

[0042] The preparation of seed liquid before fermentation comprises the following steps:

[0043] S11: Bacillus thuringiensis kurstaki was activated on a plate containing LB and cultured at 30°C for 24 h.

[0044] S12: Pick a single colony and inoculate it into a shake flask filled with shake flask culture medium, and culture it in a shaker at 30°C and 200 r / min for 24 h to form a seed solution;

[0045] S13: The seed liquid is inoculated into a seed tank filled with seed culture medium, and cultured at 30°C and 500r / min for 24h to form a pre-fermentation seed liquid; wherein the charging factor is 70%, the actual sterilization is 30min, the inoculation amount is 1wt%, and the ventilation volume is 0.6vvm. Actual sterilization refers to the actual tank sterilization, that is, the sterilization operation after adding the culture medium.

[0046] The composition of LB medium is as follows: 10 g of tryptone, 5 g of yeast extract, 10 g of sodium chloride, and 15 g of agar powder per 1000 ml of distilled water.

[0047] The shake flask medium composition is: tryptone 1wt%, yeast extract 0.5wt%, sodium chloride 1wt%, and the balance is water;

[0048] The components of the seed culture medium are: 3wt% yeast powder, 2wt% glucose, 0.2wt% potassium dihydrogen phosphate, 0.15wt% magnesium sulfate, 0.3wt% calcium carbonate, 0.03wt% sodium chloride, 0.05wt% folate, and the balance is water.

[0049] The pre-fermentation seed liquid formed in this example is used for Examples 2 to 8 and Comparative Examples 1 to 2.

[0050] Example 2

[0051] This embodiment adopts Figure 1 The fermentation tank adopts the airlift fermentation method. The volume of the inner cavity 7 of the fermentation tank is 156 tons (measured by water volume). The axis of the inner cylinder 6 is the same as the axis of the tank body 1. The diameter of the inner cylinder 6 is 4 / 5 of the inner diameter of the tank body 1, and the height is 0.7 of the height of the tank body 1. The distance between the end of the other end of the horizontal distribution pipe 11 and the inner wall of the inner cylinder 6 is 1 / 16 of the inner radius of the inner cylinder 6. The distance between the outer wall of the bottom of the inner cylinder 6 and the discharge port is about 1 / 4 of the inner radius of the tank body 1. The tank body 1 is 12m high and 4m in diameter.

[0052] A gaslift fermentation method of Bacillus thuringiensis comprises the following steps:

[0053] S1, take the seed solution and fermentation medium of Example 1, and the inoculation amount is 1wt%.

[0054] S2, add the seed liquid and fermentation medium into the tank 1 from the feed port 2, turn on the heating device ( Figure 1 The filling factor is 70%, the ventilation volume is 0.7 vvm, the defoaming agent is automatically controlled to defoam, and the fermentation is carried out at 30° C. for 60 hours. The filling factor refers to the volume of the filling volume to the total volume. In this step, the pH is natural and is not controlled.

[0055] S3, put it into the tank and determine the spore quantity and protein content.

[0056] Among them, the parameters of S2 are the parameters used in the production of the existing stirred fermentation process of Bacillus thuringiensis (except for no stirring). In S2, no feeding is carried out during the fermentation culture process (in the existing stirred fermentation of Bacillus thuringiensis, no feeding is carried out during the fermentation process), and the fermentation tank culture medium is also the culture medium used in the production of the existing stirred fermentation process of Bacillus thuringiensis (that is, suitable for the existing stirred process).

[0057] The fermentation tank medium composition is:

[0058] Soy protein powder 3wt%, molasses 2wt%, potassium dihydrogen phosphate 0.2wt%, magnesium sulfate 0.15wt%, calcium carbonate 0.3wt%, sodium chloride 0.03wt%, dapoxetine 0.05wt%, and the balance is water.

[0059] The spore amount and protein content are: spore amount is 1.34x10 9 cfu / ml, protein content 1.65mg / ml.

[0060] Example 3

[0061] Compared with Example 2, the difference between this example is that the fermentation medium is different. The fermentation medium of this example is:

[0062] Yeast powder 3wt%, glucose 2wt%, potassium dihydrogen phosphate 0.2wt%, magnesium sulfate 0.15wt%, calcium carbonate 0.3wt%, sodium chloride 0.03wt%, dapoxetine 0.05wt%, and the balance is water.

[0063] The spore quantity and protein content are: spore quantity is 1.98x10 9 cfu / ml, protein content 2.03mg / ml.

[0064] Example 4

[0065] The difference between this embodiment and embodiment 3 is the ventilation volume. The ventilation volume of this embodiment is 0.6 vvm.

[0066] The spore amount and protein content are: spore amount 2.23x10 9 cfu / ml, and the protein content was 2.47 mg / ml.

[0067] Example 5

[0068] The difference between this embodiment and embodiment 3 is the ventilation volume. The ventilation volume of this embodiment is 0.65 vvm.

[0069] The spore amount and protein content are: The spore amount and protein content are: Spore amount 2.34x10 9 cfu / ml, and the protein content was 2.61 mg / ml.

[0070] Example 6

[0071] Compared with Example 5, this embodiment is different in that:

[0072] After the pH rebounds, the feeding valve 53 is opened at the same time to feed 50wt% glucose solution to control the dissolved oxygen at 20-30% and the pH at around 7 (the reason for controlling the pH is that the addition of sugar will cause the fermentation liquid to be acidic, so ammonia water needs to be added to control the pH at 7) until the tank is lowered.

[0073] The spore count is 3.14x10 9 cfu / ml, and the protein content was 3.03 mg / ml respectively.

[0074] Example 7

[0075] Compared with Example 6, this embodiment is different in that:

[0076] When the fermentation culture reaches 48 hours, the feeding valve 53 is closed to stop feeding.

[0077] The spore count is 3.87x10 9 cfu / ml, and the protein content was 3.94 mg / ml respectively.

[0078] Comparative Example 1

[0079] Compared with Example 2, the fermentation tank in this comparative example is an existing fermentation tank, and the tank body, height, diameter, volume, heating and cooling system of the fermentation tank are the same as those of this comparative example. Figure 1 The fermentation tanks are the same, and the fermentation conditions and parameters are the same, the only difference is:

[0080] 1. The fermentation tank is equipped with a stirrer, but not with Figure 1 The inner cylinder 6 and the supporting structures such as the first pushing unit 10.

[0081] 2. Stirring speed: 250r / min.

[0082] The spore amount and protein content of this comparative example are as follows: the spore amount is 3.55x10 9 cfu / ml, protein content 3.78mg / ml.

[0083] Comparative Example 2

[0084] Compared with Comparative Example 1, this comparative example has the following differences:

[0085] After the pH rebounds, start adding 50% glucose solution, control the dissolved oxygen at 20-30%, and control the pH at around 7 (the reason for controlling the pH is that the addition of sugar will cause the fermentation liquid to be acidic, so ammonia water needs to be added to control the pH at 7) until the tank is placed.

[0086] The spore amount and protein content of this comparative example are as follows: the spore amount is 3.50x10 9 cfu / ml, protein content 3.70mg / ml.

[0087] From the above-mentioned examples 2 to 7, and comparative examples 1 to 2, compared with comparative example 1, Example 2 reduces the effect of the fermented liquid while saving energy consumption. Compared with Example 2, Example 3 shows that the fermentation medium of Example 3 is more suitable for the airlift fermentation process of Bacillus thuringiensis. Compared with the examples, the ventilation volume of Example 4 and Example 5 is more suitable for the airlift fermentation process of Bacillus thuringiensis than 0.6vvm or 0.7vvm. Compared with Example 6, Example 5, as well as Comparative Examples 1 and 2, for the fermentation of Bacillus thuringiensis, feeding does not improve the fermentation liquid effect of stirred fermentation, but significantly improves the fermentation liquid effect of airlift. Compared with Example 6, Example 7 shows that the effect of stopping feeding after feeding for 48h of fermentation is significantly better than feeding for 60h, and finally completely realizes that airlift fermentation replaces stirred fermentation, and obtains a fermented liquid with better effect than the existing one by controlling the feeding time.

[0088] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. For those skilled in the art, the present invention may have various modifications and variations. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present invention shall be included in the protection scope of the present invention.

Claims

1. A method for airlift fermentation of Bacillus thuringiensis without stirring during the fermentation process, characterized in that: The following steps are involved: S1, taking Bacillus thuringiensis seed solution and fermentation medium, the inoculation amount is 0.5-10wt%; S2, adding seed liquid and fermentation medium into an airlift fermenter, with a loading factor of 60-80%, aeration of 0.6-0.8 vvm, automatic defoaming control by defoaming agent, pH 6.5-7.5, and fermentation culture at 28-31°C for 50-70 hours; S3, the Bacillus thuringiensis fermentation liquid is obtained in the lower tank.

2. The airlift fermentation method of Bacillus thuringiensis according to claim 1, characterized in that: In S2, when the fermentation culture reaches pH rebound, 30-60 wt % glucose solution is continuously added to control the dissolved oxygen at 20-30%.

3. The method according to claim 2, characterized in that: In S2, 50 wt% glucose solution is added until 5 to 15 hours from the end of the fermentation, and then the addition of 30 to 60 wt% glucose solution is stopped.

4. The airlift fermentation method of Bacillus thuringiensis according to any one of claims 1 to 3, characterized in that: The fermentation medium comprises: Yeast powder 1-10wt%, glucose 1-5wt%, potassium dihydrogen phosphate 0.1-0.5wt%, magnesium sulfate 0.08-0.2wt%, calcium carbonate 0.1-0.5wt%, sodium chloride 0.01-0.05wt%, dapoxetine 0.02-0.08wt%, and the balance is water.

5. The airlift fermentation method of Bacillus thuringiensis according to any one of claims 1 to 3, characterized in that: The fermentation medium comprises: 1-10wt% soy protein powder, 1-5wt% molasses, 0.1-0.5wt% potassium dihydrogen phosphate, 0.08-0.2wt% magnesium sulfate, 0.1-0.5wt% calcium carbonate, 0.01-0.05wt% sodium chloride, 0.02-0.08wt% folate, and the balance is water.

6. The airlift fermentation method of Bacillus thuringiensis according to any one of claims 1 to 5, characterized in that: In S1, the Bacillus thuringiensis seed solution is mainly prepared by the following steps: S11, Bacillus thuringiensis kurstaki was activated on plates containing LB medium and cultured at 28-31°C for 12-36 h; S12, picking a single colony and inoculating it into a shake flask filled with shake flask culture medium, and culturing it in a shaker at 28-31°C and 100-500 r / min for 12-36 hours to form a seed solution; S13, the seed liquid is inoculated into a seed tank filled with a seed tank culture medium, and cultured at 28-31°C and 100-500r / min for 12-36h to form a pre-fermentation seed liquid, with a charging factor of 60-80%, an actual digestion time of 20-50min, an inoculation amount of 0.5-10wt%, and an aeration volume of 0.6-0.7vvm.

7. The airlift fermentation method of Bacillus thuringiensis according to any one of claims 1 to 6, characterized in that: In S11, the components of LB culture medium are: 5-15g of tryptone, 2-8g of yeast extract, 5-15g of sodium chloride, and 10-20g of agar powder per 1000ml of distilled water; in S12, the components of shake flask culture medium are: 0.5-1.5wt% of tryptone, 0.2-0.8wt% of yeast extract, 0.5-1.5wt% of sodium chloride, and the balance is water; in S13, the components of seed culture medium are: 1-5wt% of yeast powder, 1-3wt% of glucose, 0.1-0.3wt% of potassium dihydrogen phosphate, 0.1-0.2wt% of magnesium sulfate, 0.2-0.4wt% of calcium carbonate, 0.02-0.04wt% of sodium chloride, 0.02-0.08wt% of folate, and the balance is water.

8. The airlift fermentation method of Bacillus thuringiensis according to any one of claims 1 to 7, characterized in that: The Bacillus thuringiensis is Bacillus thuringiensis kurstaki.

9. The airlift fermentation method of Bacillus thuringiensis according to any one of claims 1 to 8, characterized in that: The airlift fermentation tank comprises a tank body, on which a feed inlet, a discharge inlet, and a feed and air passage are provided, the tank body forms an inner cavity of the tank body, the feed inlet is provided at the top of the tank body, the discharge inlet is provided at the bottom of the tank body, the feed and air passages are located at the lower part of the tank body, the feed and air passages are connected with a main pipe, the main pipe passes through the tank body, a part of the main pipe is located in the inner cavity of the tank body, and the rest of the main pipe is located outside the inner cavity of the tank body, the end of the main pipe located outside the inner cavity of the tank body is connected with an air inlet pipe and a feed pipe, a feed valve and a feed cup are installed on the feed pipe, an air inlet valve is installed on the air inlet pipe, an inner cylinder is installed in the inner cavity of the tank body, the top of the inner cylinder is provided with an upper opening, the bottom is provided with a lower opening, the inner cylinder forms an inner cavity of the inner cylinder, the main pipe is located in the tank body Part of the inner cavity passes through the inner cylinder and is located in the inner cavity of the inner cylinder. Its end opening is downward and is connected to the distributor. A first pushing unit is arranged below the bottom of the distributor. More than three horizontal distribution pipes are connected to the side wall of the distributor. The more than three horizontal distribution pipes are evenly distributed along the circumference. More than two second pushing units are arranged above each horizontal distribution pipe. The first pushing unit is located below the bottom of the inner cylinder, and the second pushing unit is located in the inner cavity of the inner cylinder. The first pushing unit is connected to the distributor through a connecting pipe, and the second pushing unit is connected to the horizontal distribution pipe through a connecting pipe. The pushing inlet of the first pushing unit is at the top and the pushing outlet is at the bottom, and the pushing inlet of the second pushing unit is at the bottom and the pushing outlet is at the top.

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