Method and system for high-throughput screening of flavone transforming bacteria

Abstract

The invention relates to the technical field of microbial screening, in particular to a method for high-throughput screening of flavone-transformed bacteria and a screening system thereof. The method comprises the following steps: firstly, inoculating a to-be-detected strain into a basal culture medium containing a flavone substrate for fermentation, then scanning fermentation broth at two different time points by using a full-wavelength microplate reader through a quartz microplate, calculating an absorbance difference value at at least one preset wavelength, and finally, calculating the absorbance difference value at least one preset wavelength. The strains with the absorbance difference larger than a preset threshold value are screened as primary screening positive strains. Compared with the traditional technology, the method has the advantages that the screening efficiency is improved by hundreds of times, the sample consumption is reduced by 75%, the industrial bottleneck that multi-structure flavonoids need to be repeatedly developed is broken through, and an innovative tool is provided for microbial resource development.

Description

TECHNICAL FIELD

[0001] The present application belongs to the technical field of microorganism screening, and particularly relates to a method for high-throughput screening of flavone conversion bacteria and a screening system thereof. BACKGROUND

[0002] Flavone conversion bacteria refer to a class of microorganism strains capable of modifying flavonoids (such as hydroxylation, glycosylation, ring cleavage, etc.) through their own metabolic enzyme system, and the conversion products thereof often have enhanced biological activity, solubility or new pharmacological functions. Due to the differentiated application values of flavones of different structural types (such as isoflavones like isobavone, dihydroflavones like tangeritin, isoflavones like daidzein, flavonols like icariin, isoflavans like glabridin, etc.) in the fields of drug development (such as anti-tumor, anti-oxidation, cardiovascular protection) and functional foods, screening of strains with broad-spectrum or specific conversion ability has become a core link for the development of high-value flavone derivatives.

[0003] Currently, the industry mainly relies on two types of traditional screening technologies: high-performance liquid chromatography (HPLC): the fermentation broth of each strain needs to be individually subjected to chromatographic separation, and the activity of the strain is judged by comparing the changes in characteristic peaks before and after conversion. Although this method is highly accurate, it takes 30-60 minutes for single sample detection, and it takes several weeks for thousands of strains screening, and it consumes ≥2 mL of fermentation broth sample, which is difficult to handle microstrains. Ultraviolet spectrophotometry: the conversion efficiency is indirectly evaluated by the absorbance change at a specific wavelength (such as 280 nm). Although it is faster than HPLC, it cannot distinguish between structurally similar flavones and their metabolites, with a mis-screening rate of more than 40%, and different flavone types need to be independently optimized for detection wavelength (such as 256 nm for daidzein and 270 nm for icariin), and the method has poor universality.

[0004] Therefore, it is of great significance to develop a universal strain screening method that can simultaneously cover multiple flavone structural types, support rapid preliminary screening of microsamples and improve the throughput by more than 100 times, to meet the needs of industrial strain development. SUMMARY

[0005] The present application aims to overcome the problems of slow screening speed, large sample consumption, low accuracy and inability to simultaneously detect multiple flavone structural types in high throughput in the prior art method for screening conversion bacteria of different flavone structural types, and to provide a method for high-throughput screening of flavone conversion bacteria and a screening system thereof. The method can simultaneously cover multiple flavone structural types, support rapid preliminary screening of microsamples and improve the throughput by more than 100 times, to meet the needs of industrial strain development, which is of great significance.

[0006] To achieve the above-mentioned objectives, the present application adopts the following technical solutions:

[0007] The first aspect of this invention discloses a method for high-throughput screening of flavonoid-transforming bacteria, comprising:

[0008] Step 1: Inoculate the test strain into a basic medium containing a flavonoid substrate for fermentation. The flavonoid substrate is at least one of the following structural types: chalcones, dihydroflavones, isoflavones, flavonols, and isoflavones.

[0009] Step 2: At at least two different time points during fermentation, use a full-wavelength microplate reader to scan the fermentation broth at a full wavelength of 200-500 nm through a quartz microplate.

[0010] Step 3: Based on the scanning results obtained in Step 2, calculate the absorbance difference at at least one preset wavelength;

[0011] Step 4: Select strains with absorbance difference greater than the preset threshold as initial positive strains.

[0012] The high-throughput screening method for flavonoid-transforming bacteria provided by this invention first involves inoculating the test strain into a basal culture medium containing flavonoid substrate for fermentation. Then, the fermentation broth at two different time points is scanned using a full-wavelength microplate reader, and the absorbance difference at at least one preset wavelength is calculated. Finally, strains with absorbance differences greater than a preset threshold are selected as initial positive strains. Compared to traditional techniques, this method improves screening efficiency by a hundredfold, reduces sample consumption by 75%, and overcomes the industry bottleneck of requiring repeated development of multi-structured flavonoids, providing an innovative tool for microbial resource development.

[0013] Furthermore, the basal culture medium contains: ammonium nitrate 1.8-2.2 g / L, magnesium sulfate 0.4-0.6 g / L, dipotassium hydrogen phosphate 0.8-1.2 g / L, pH 6.5-7.5.

[0014] Furthermore, the concentration of the flavonoid substrate in the basal culture medium is 0.01-1 mg / mL.

[0015] Furthermore, the flavonoid substrate is selected from at least one of glycyrrhizin, isoliquiritin, daidzein, icariin, hesperidin, and phloretin.

[0016] Furthermore, at least two distinct time points are included, including the fermentation start point and the fermentation end point.

[0017] Furthermore, the preset wavelength mentioned in step 3 is selected according to the flavonoid structure type as at least one of 281±5nm, 316±5nm, 350±5nm and 370±5nm.

[0018] Furthermore, the preset threshold mentioned in step 4 is 0.15-0.3.

[0019] Furthermore, step 5 involves validating the fermentation broth of the initially screened positive strains using high-performance liquid chromatography (HPLC). This method combines the high throughput and rapid detection capabilities of a full-wavelength microplate reader with the high resolution and accurate quantification advantages of HPLC, overcoming the shortcomings of traditional methods. The full-wavelength microplate reader can complete the initial screening of a large number of strains in a short time, significantly narrowing the range of samples to be tested. HPLC then performs precise qualitative and quantitative analysis on a small number of key strains. This two-step method significantly improves screening efficiency and accuracy, bringing technological innovation to the screening of microbial transformation strains.

[0020] A second aspect of the present invention provides a flavonoid-transforming bacteria screening system for implementing the above-described method, comprising:

[0021] A full-wavelength microplate reader, configured to perform absorbance scanning in the wavelength range of 200-500 nm;

[0022] Quartz microplates are used to hold fermentation broth samples containing bacterial strains, flavonoid substrates, and basal culture medium for testing.

[0023] The data processing unit, communicatively connected to the full-wavelength microplate reader, is configured as follows:

[0024] a. Receives and stores full-wavelength absorbance scan data at different time points;

[0025] b. Calculate the absorbance difference at at least one preset wavelength;

[0026] c compares the calculated absorbance difference with a preset threshold.

[0027] d outputs or identifies the strain information corresponding to samples whose absorbance difference is greater than a preset threshold.

[0028] A third aspect of this invention provides a special culture medium for screening flavonoid-transforming bacteria, comprising the following components:

[0029] Basic culture medium: ammonium nitrate 1.8-2.2 g / L, magnesium sulfate 0.4-0.6 g / L, dipotassium hydrogen phosphate 0.8-1.2 g / L, pH 6.5-7.5;

[0030] Flavonoid substrates: glycyrrhizin, isoliquiritin, daidzein, icariin, hesperidin or phloretin, at a concentration of 0.01-1 mg / mL.

[0031] The culture medium provided by this invention can significantly reduce the UV background interference of non-target flavonoids, which helps to improve the accuracy of the screening method of this application.

[0032] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0033] 1. The high-throughput screening method for flavonoid-transforming bacteria provided by this invention first involves inoculating the test strain into a basal culture medium containing flavonoid substrate for fermentation. Then, the fermentation broth at two different time points is scanned using a full-wavelength microplate reader, and the absorbance difference at at least one preset wavelength is calculated. Finally, strains with absorbance differences greater than a preset threshold are selected as initial positive strains. Compared to traditional techniques, this method improves screening efficiency by a hundredfold, reduces sample consumption by 75%, and overcomes the industry bottleneck of repeated development of multi-structure flavonoids, providing an innovative tool for microbial resource development.

[0034] 2. The method of this invention is simple to operate, reacts rapidly, and has a short operation time. It has no requirements for experimental time or temperature, and requires very little sample (less than 0.5 mL). It can be used for the rapid detection of changes in the content of glycyrrhizin, isoliquiritigenin, daidzein, icariin, hesperidin, and phlorizin in fermentation broth, as well as whether structural transformation has occurred. Attached Figure Description

[0035] Figure 1 This is a specific data diagram for an example.

[0036] Figure 2 This is a graph showing data from a full-wavelength microplate reader used to screen transformed strains, as an example.

[0037] Figure 3 The image shows the HPLC verification results for the example.

[0038] Figure 4 The image shows the LC-QTOF results of the mixture of the strain fermentation broth and daidzein solution in the example. Detailed Implementation

[0039] The present invention will now be described in further detail with reference to specific embodiments. However, this should not be construed as limiting the scope of the present invention to the following embodiments; all technologies implemented based on the content of the present invention fall within the scope of the present invention.

[0040] The first aspect of this embodiment discloses a method for high-throughput screening of flavonoid-transforming bacteria, comprising:

[0041] Step 1: Inoculate the test strain into a basic medium containing a flavonoid substrate for fermentation. The flavonoid substrate is at least one of the following structural types: chalcones, dihydroflavones, isoflavones, flavonols, and isoflavones.

[0042] Step 2: At at least two different time points during fermentation, use a full-wavelength microplate reader to scan the fermentation broth at a full wavelength of 200-500 nm through a quartz microplate.

[0043] Step 3: Based on the scanning results obtained in Step 2, calculate the absorbance difference at at least one preset wavelength;

[0044] Step 4: Select strains with absorbance difference greater than the preset threshold as initial positive strains.

[0045] Compared to traditional techniques, this method improves screening efficiency by a hundredfold, reduces sample consumption by 75%, and breaks through the industry bottleneck of repeated development of multi-structure flavonoids, providing an innovative tool for the development of microbial resources.

[0046] Traditional methods for screening flavonoid-transforming strains are inefficient, have low throughput, and are time-consuming. This embodiment provides an innovative solution for screening transforming strains using a full-wavelength microplate reader. The principle is based on the fact that the full-wavelength microplate reader uses light emitted from a light source, which is filtered or grating into monochromatic light to illuminate the sample in the microplate. Molecules in the sample absorb specific wavelengths of light, producing different absorbance values. The microplate reader detects changes in absorbance values ​​to reflect the dynamic changes in sample composition. Different flavonoid substrates, such as chalcone and dihydroflavonoids, are added. After fermentation, the full-wavelength microplate reader continuously monitors and records the changes in absorbance values ​​across the entire wavelength range of the fermentation system. Different strains have different metabolic transformation capabilities, leading to differences in the optical properties of the fermentation system and thus exhibiting different light absorption curves. By analyzing these curves, strains with transformation potential can be quickly identified. Subsequent high-performance liquid chromatography (HPLC) is used to detect these potential strains. Precise analysis of the changes in the composition and content of flavonoids before and after fermentation further confirms the strain's transformation ability and product activity.

[0047] In some embodiments, the basal culture medium comprises: ammonium nitrate 1.8-2.2 g / L, magnesium sulfate 0.4-0.6 g / L, dipotassium hydrogen phosphate 0.8-1.2 g / L, pH 6.5-7.5. This culture medium can significantly reduce UV background interference from non-target flavonoids, which helps to improve the accuracy of the screening method of this application.

[0048] In some embodiments, the concentration of the flavonoid substrate in the basal culture medium is 0.01-1 mg / mL.

[0049] In some embodiments, the flavonoid substrate is selected from at least one of glycyrrhizin, isoliquiritin, daidzein, icariin, hesperidin, and phloretin.

[0050] In some embodiments, at least two different time points include the fermentation start point and the fermentation end point.

[0051] In some embodiments, the preset wavelength in step 3 is selected according to the flavonoid structure type as at least one of 281±5nm, 316±5nm, 350±5nm, and 370±5nm.

[0052] In some embodiments, the preset threshold in step 4 is 0.15-0.3. For example, the preset threshold in step 4 is 0.15, 0.2, 0.25, or 0.3. Controlling an appropriate preset threshold results in higher screening accuracy.

[0053] In some embodiments, the method further includes: step 5, validating the fermentation broth of the initially screened positive strains using high-performance liquid chromatography (HPLC). This method combines the high throughput and rapid detection characteristics of a full-wavelength microplate reader with the high resolution and accurate quantification advantages of HPLC, overcoming the shortcomings of traditional methods. A full-wavelength microplate reader can complete the initial screening of a large number of strains in a short time, significantly narrowing the range of samples to be tested. HPLC then performs precise qualitative and quantitative analysis on a small number of key strains. This two-step method significantly improves screening efficiency and accuracy, bringing technological innovation to the screening of microbial transformation strains.

[0054] The second aspect of this embodiment provides a flavonoid-transforming bacteria screening system for implementing the above method, including:

[0055] A full-wavelength microplate reader, configured to perform absorbance scanning in the wavelength range of 200-500 nm;

[0056] Quartz microplates are used to hold fermentation broth samples containing bacterial strains, flavonoid substrates, and basal culture medium for testing.

[0057] The data processing unit, communicatively connected to the full-wavelength microplate reader, is configured as follows:

[0058] a. Receives and stores full-wavelength absorbance scan data at different time points;

[0059] b. Calculate the absorbance difference at at least one preset wavelength;

[0060] c compares the calculated absorbance difference with a preset threshold.

[0061] d outputs or identifies the strain information corresponding to samples whose absorbance difference is greater than a preset threshold.

[0062] The third aspect of this embodiment provides a special culture medium for screening flavonoid-transforming bacteria, which consists of the following:

[0063] Basic culture medium: ammonium nitrate 1.8-2.2 g / L, magnesium sulfate 0.4-0.6 g / L, dipotassium hydrogen phosphate 0.8-1.2 g / L, pH 6.5-7.5;

[0064] Flavonoid substrates: glycyrrhizin, isoliquiritin, daidzein, icariin, hesperidin or phloretin, at a concentration of 0.01-1 mg / mL.

[0065] The culture medium provided in this embodiment can significantly reduce the UV background interference of non-target flavonoids, which helps to improve the accuracy of the screening method of this application.

[0066] To better understand the technical solutions of the above embodiments, the following detailed embodiments are provided for further explanation.

[0067] Example 1

[0068] Step 1: The linearity and effects of basal culture medium on chalcone isoliquiritigenin, dihydroflavonoid hesperidin, isoflavones soy isoflavones, flavonols icariin, isoflavones glycyrrhizin, and dihydroflavonoid phlorizin. The basal culture medium significantly reduced the UV background interference of non-target flavonoids.

[0069] Icariin Standard Solution

[0070] Take 1g of icariin standard, dissolve it in a small amount of methanol, transfer it to a 1000mL volumetric flask, rinse the beaker with methanol, add the flask, and bring the volume to 1000mL with methanol. Prepare a 1mg / mL icariin stock solution.

[0071] Eleven serial standard solutions were prepared using methanol from a 1 mg / mL icariin stock solution. Take 0.1 mL, 0.01 mL, 0.02 mL, 0.03 mL, 0.04 mL, 0.05 mL, 0.06 mL, 0.07 mL, 0.6 mL, and 0.8 mL of the icariin stock solution, respectively, and add 1 mL to basal culture medium to a final volume of 1 mL to prepare standard solutions with concentrations of 0.1 mg / mL, 0.01 mg / mL, 0.02 mg / mL, 0.03 mg / mL, 0.04 mg / mL, 0.04 mg / mL, 0.05 mg / mL, 0.06 mL, 0.07 mL, 0.6 mL, and 0.04 mL. The concentration range of icariin standard solutions was 0.01 mg / mL to 1 mg / mL. A linear regression equation was established using the absorbance along the Y-axis for these 11 concentrations. The linear regression equation for icariin standard solutions at 270 nm for concentrations of 0.01–0.4 mg / mL was t = 0.0052x + 0.0734R. 2 =0.9991, the linear regression equation for icariin standard solution at 270 nm in the range of 0.01-0.05 mg / mL is t = 0.0191x + 0.0274R. 2 =0.9921, the linear regression equation of icariin standard solution at 0.05-1 mg / mL at 270 nm is y = 0.0038x + 0.7125R. 2 The limit of detection for icariin using the full-wavelength ELISA reader was 0.015 μg / mL, and the limit of quantification was 0.108 μg / mL. The relative standard deviation of icariin was calculated to be 1.2% using Table 1, indicating that icariin showed good repeatability under the full-wavelength ELISA reader.

[0072] Standard solutions of daidzein, hesperidin, and phlorizin

[0073] Linear regression equations were established using the absorbance along the Y-axis for 11 concentrations of daidzein standard solutions ranging from 0.01 to 0.4 mg / mL at 256 nm. The linear regression equation was t = 0.0004x + 0.0812R. 2 =0.9502. The linear regression equation for daidzein standard solution at 256 nm in the range of 0.01-0.05 mg / mL is t = 0.0397x + 0.0208R² = 0.9966. The limit of detection for daidzein by the full-wavelength ELISA reader is 0.118 μg / mL, and the limit of quantitation is 0.213 μg / mL. The relative standard deviation of daidzein can be calculated as 0.206% from Table 2, indicating that daidzein has good repeatability under the full-wavelength ELISA reader.

[0074] The linear regression equation for hesperidin standard solutions at 283 nm in the range of 0.01–0.4 mg / mL is y = 0.009x + 0.1136R. 2 =0.9971. The linear regression equation for daidzein standard solution at 283 nm in the range of 0.01-0.05 mg / mL is yy = 0.0482x + 0.1353R. 2 =0.0358, the detection limit of the full-wavelength ELISA reader for daidzein is 0.210 μg / mL, and the quantitation limit is 0.346 μg / mL. The relative standard deviation of daidzein can be calculated from Table 3 as 0.48%, indicating that hesperidin has good repeatability under the full-wavelength ELISA reader.

[0075] The linear regression equation for phlorizin standard solutions at 285 nm in the range of 0.01–0.4 mg / mL is y = 0.0188x + 0.0204R. 2 =0.9927. The linear regression equation for phlorizin standard solutions at 285 nm in the range of 0.01-0.05 mg / mL is yy = 0.0191x + 0.0247R. 2 =0.9921, the detection limit of phlorizin by the full-wavelength ELISA reader is 0.102 μg / mL, and the quantification limit is 0.148 μg / mL. The relative standard deviation of daidzein can be calculated from Table 4 as 0.448%, indicating that phlorizin has good repeatability under the detection of the full-wavelength ELISA reader.

[0076] Table 1. Icariin

[0077]

[0078] Table 2. Daidzein

[0079]

[0080] Table 3 Hesperidin

[0081]

[0082] Table 4 Phloretin

[0083]

[0084] A standard solution of 0.01 mg / mL glycyrrhizin, isoliquiritigenin, daidzein, icariin, hesperidin, and phlorizin was selected at a concentration of 300 μL. A 300 μL basal culture medium was used, with 300 μL methanol as a blank control. The solutions were added to 96-well quartz microplates and analyzed at 200-500 nm using a full-wavelength microplate reader. The absorbance of the basal culture medium and the blank methanol was lower than that of the glycyrrhizin standard solution before 300 nm, but had no effect on the absorbance of the glycyrrhizin, isoliquiritigenin, daidzein, icariin, hesperidin, and phlorizin standard solutions. The x-axis represents wavelength, and the y-axis represents absorbance. Figure 1 .

[0085] The full-wavelength microplate reader used in the experiment was an Epoch model, and the 96-well quartz plate was made of ultraviolet optical quartz glass that could detect substances at wavelengths of 220-2500nm.

[0086] Repeatability tests show that, using the method of this invention and following the specific implementation steps, the contents of glycyrrhizin, isoliquiritin, daidzein, icariin, hesperidin, and phlorizin in randomly selected test samples can be accurately determined at appropriate concentrations. The results of the repeatability tests demonstrate that the method of this invention is analytically stable and meets the repeatability requirements.

[0087] Step Two: After successful fermentation of any strain, centrifuge at 10,000 rpm for 10 minutes to collect the precipitated bacterial cells. Add these cells to a basal culture medium containing glycyrrhizin, isoliquiritin, daidzein, icariin, hesperidin, and phloretin. After co-culturing for 0 hours and 168 hours, use a full-wavelength microplate reader to collect the flavonoid-containing fermentation broth at a wavelength of 200-500 nm for full-wavelength detection. For example... Figure 2 The fermentation broth of strains with differences of 0h and blank (basal culture medium with flavonoid standards) at 281nm, 370nm, 350nm, and 316nm greater than 0.2 was subjected to HPLC analysis. Figure 3 The HPLC results of some transformed strains show that a total of two strains that can transform glycyrrhizin, 18 strains that can transform isoliquiritin, one strain that can transform daidzein, four strains that can transform icariin, one strain that can transform hesperidin, and two strains that can transform phloretin were screened.

[0088] Step 3: 1500 bacterial strains were screened using a full-wavelength microplate reader. 50 strains with an OD difference greater than 0.2 were then analyzed by HPLC. It was found that 20 of these 50 strains showed small peaks next to the standard peaks of glycyrrhizin, isoliquiritin, daidzein, icariin, hesperidin, and phlorizin in HPLC, demonstrating the accuracy and speed of the method in screening large quantities of flavonoids that can be converted into different structures. These were presumed to be flavonoid products, and their structures were further identified by high-resolution mass spectrometry. Figure 4 LC-QTOF results of the mixture of fermentation broth of the strain and daidzein solution.

[0089] Based on this invention, standard solutions of glycyrrhizin, isoliquiritigenin, daidzein, icariin, hesperidin, and phlorizin were added to a basic culture medium, and endophytic plant strains were added for fermentation. The full-wavelength curves at 0h and 168h after fermentation were detected using a full-wavelength microplate reader. By comparing the full-wavelength change curves, the selected strains were verified by high-performance liquid chromatography (HPLC) to identify characteristic peaks of the transformation products, such as... Figure 3 Then, LC-QTOF is performed to obtain the structure, such as Figure 4 The results of screening for a strain LC-QTOF capable of converting daidzein are shown. The 16rsDNA sequence of the screened strain is shown in SEQ ID No. 1.

[0090] Sequence SEQ ID No.1:

[0091]

[0092] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, and improvements made within the spirit and principles of the present invention should be included within the protection scope of the present invention.

Claims

1. A method for high-throughput screening of flavonoid-transforming bacteria, characterized in that, include: Step 1: Inoculate the test strain into a basic medium containing a flavonoid substrate for fermentation. The flavonoid substrate is at least one of the following structural types: chalcones, dihydroflavones, isoflavones, flavonols, and isoflavones. Step 2: At at least two different time points during fermentation, use a full-wavelength microplate reader to scan the fermentation broth at a full wavelength of 200-500 nm through a quartz microplate. Step 3: Based on the scanning results obtained in Step 2, calculate the absorbance difference at at least one preset wavelength; Step 4: Select strains with absorbance differences greater than a preset threshold as initial positive strains.

2. The method according to claim 1, characterized in that, The basal culture medium contains: ammonium nitrate 1.8-2.2 g / L, magnesium sulfate 0.4-0.6 g / L, dipotassium hydrogen phosphate 0.8-1.2 g / L, pH 6.5-7.

5.

3. The method according to claim 1, characterized in that, The concentration of the flavonoid substrate in the basal culture medium was 0.01-1 mg / mL.

4. The method according to claim 1, characterized in that, The flavonoid substrate is selected from at least one of glycyrrhizin, isoliquiritin, daidzein, icariin, hesperidin, and phloretin.

5. The method according to claim 1, characterized in that, At least two distinct time points, including the fermentation start point and the fermentation end point.

6. The method according to claim 1, characterized in that, The preset wavelength mentioned in step 3 is selected according to the flavonoid structure type as at least one of 281±5nm, 316±5nm, 350±5nm and 370±5nm.

7. The method according to claim 1, characterized in that, The preset threshold mentioned in step 4 is 0.15-0.

3.

8. The method according to any one of claims 1-7, characterized in that, Also includes: Step 5: Verify the fermentation broth of the initially screened positive strains by high performance liquid chromatography.

9. A flavonoid-transforming bacteria screening system, used to implement the method according to any one of claims 1-8, characterized in that, include: A full-wavelength microplate reader, configured to perform absorbance scanning in the wavelength range of 200-500 nm; Quartz microplates are used to hold fermentation broth samples containing bacterial strains, flavonoid substrates, and basal culture medium for testing. The data processing unit, communicatively connected to the full-wavelength microplate reader, is configured as follows: a. Receives and stores full-wavelength absorbance scan data at different time points; b. Calculate the absorbance difference at at least one preset wavelength; c compares the calculated absorbance difference with a preset threshold. d outputs or identifies the strain information corresponding to samples whose absorbance difference is greater than a preset threshold.

10. A special culture medium for screening flavonoid-transforming bacteria, characterized in that, It consists of the following: Basic culture medium: ammonium nitrate 1.8-2.2 g / L, magnesium sulfate 0.4-0.6 g / L, dipotassium hydrogen phosphate 0.8-1.2 g / L, pH 6.5-7.5; Flavonoid substrates: glycyrrhizin, isoliquiritin, daidzein, icariin, hesperidin or phloretin, at a concentration of 0.01-1 mg / mL.

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