CN110846234A - Pleurotus cornucopiae culture medium - Google Patents
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Abstract
The invention discloses a pleurotus cornucopiae culture medium. The culture medium comprises a solid culture medium and a liquid culture medium, wherein the solid culture medium comprises the following components: 200g of potato, 15g of agar, 10g of glucose and 10-20 g of mannitol, and adding distilled water to a constant volume of 1L; the composition of the liquid medium was: 200g of potato, 10g of glucose, 10-20 g of mannitol and distilled water to a constant volume of 1L. The preparation method of the culture medium is simple, good nutrient substances and space environment can be provided for the growth of the pleurotus cornucopiae, the growth speed of pleurotus cornucopiae mycelia is remarkably increased, the biomass of the mycelia is increased, the laccase activity of the mycelia is improved, the cost is low, and the pleurotus cornucopiae growth vigor is excellent.
Description
Technical Field
The invention belongs to the technical field of pleurotus cornucopiae cultivation, and relates to a pleurotus cornucopiae culture medium.
Background
Pleurotus cornucopiae (Paul. Pers.) Rolland) is called Pleurotus cornucopiae, Pleurotus cornucopiae. The pleurotus cornucopiae is rich in nutritional ingredients such as proteins, various vitamins and minerals, is delicious in taste, and has the effects of resisting tumors, reducing blood pressure, improving metabolism, enhancing immunity and the like. Chinese patent 201210023901 discloses a liquid fermentation culture method of Pleurotus cornucopiae strain, and studies the influence of different carbon sources on the growth of Pleurotus cornucopiae, and finds that Pleurotus cornucopiae grows best in a culture medium with mannitol as the carbon source, the rest is fructose, glucose, sucrose and soluble starch in turn, the utilization rate of two monosaccharides of mannitol and fructose is good, and the optimal culture conditions are 3% of mannitol, 1% of yeast extract and 0.2% of KH2PO4And 0.05% MgSO4·7H2And O. However, the culture medium is added with mannitol, nitrogen source yeast extract and corn flour, and is subjected to fermentation culture.
Disclosure of Invention
The invention aims to provide a simple pleurotus cornucopiae culture medium which can remarkably accelerate the growth speed of pleurotus cornucopiae hyphae and improve the biomass of the pleurotus cornucopiae hyphae and the laccase activity.
The technical scheme for realizing the purpose of the invention is as follows:
the pleurotus cornucopiae culture medium comprises a solid culture medium and a liquid culture medium, wherein the solid culture medium comprises the following components: 200g of potato, 15g of agar, 10g of glucose and 10-20 g of mannitol, and adding distilled water to a constant volume of 1L; the composition of the liquid medium was: 200g of potato, 10g of glucose, 10-20 g of mannitol and distilled water to a constant volume of 1L.
The solid culture medium and the liquid culture medium are sterilized at 121 ℃ for 20min, and are cooled for inoculation.
The inoculation is carried out on an ultra-clean workbench which is subjected to ultraviolet sterilization for 30 min.
The inoculation amount of the liquid culture medium is 10%.
The invention also provides a culture method of the pleurotus cornucopiae, which comprises the following specific steps:
inoculating Pleurotus cornucopiae mycelium into the above solid culture medium or liquid culture medium, and culturing at 25 deg.C in dark.
Preferably, the culture is carried out in a constant temperature shaker at 25 ℃ at a rotation speed of 150rpm while being inoculated in a liquid medium.
Preferably, when inoculated in a solid medium, the culture is carried out in an incubator at a constant temperature of 25 ℃.
Compared with the prior art, the invention has the following advantages:
the culture medium can provide good nutrient substances and a space environment for growth of the pleurotus cornucopiae, the preparation method is simple, mannitol is only added into a conventional glucose culture medium, the addition amount of the glucose is controlled to be 1% and the addition amount of the mannitol is controlled to be 1-2%, and when the mass ratio of the glucose to the mannitol is 1:2, the growth speed of pleurotus cornucopiae mycelia can be remarkably accelerated, the biomass of the mycelia is increased, the laccase activity of the mycelia is improved, the cost is low, and the growth vigor of the pleurotus cornucopiae is excellent.
Drawings
FIG. 1 is a graph showing the growth of the hyphae of Pleurotus cornucopiae of examples 1 and 2 and comparative examples 1 and 2 during the culture of Pleurotus cornucopiae;
FIG. 2 is a graph showing the growth rate of the hyphae of Pleurotus cornucopiae of examples 1 and 2 and comparative examples 1 and 2 during the solid culture of Pleurotus cornucopiae;
FIG. 3 is a graph showing the dry weight of Pleurotus cornucopiae mycelia of examples 1 and 2 and comparative examples 1 and 2 during the liquid culture of Pleurotus cornucopiae;
FIG. 4 is a graph showing laccase activity of Agaricus blazei Murill mycelia of examples 1 and 2 and comparative examples 1 and 2 obtained by liquid culture of Pleurotus cornucopiae;
FIG. 5 is a protein electrophoresis chart of Agaricus blazei Murill mycelia in examples 1 and 2 and comparative examples 1 and 2 obtained by liquid culture of Agaricus blazei Murill.
Detailed Description
The invention is further illustrated below with reference to specific embodiments and the accompanying drawings. It should be understood that these examples are for illustrative purposes only and are not intended to limit the scope of the present invention. Further, it should be understood that various changes or modifications of the present invention may be made by those skilled in the art after reading the teaching of the present invention, and such equivalents may fall within the scope of the present invention as defined in the appended claims.
Comparative example 1
The common culture medium for the pleurotus cornucopiae comprises a solid culture medium and a liquid culture medium, wherein the solid culture medium comprises the following components: 200g of potato, 15g of agar and 20g of glucose, distilled water is added to the mixture to be constant volume to 1L, and the composition of a liquid culture medium is as follows: 200g of potato and 20g of glucose, adding distilled water to a constant volume of 1L, and using a 250mL triangular flask containing 100mL of liquid culture medium per bottle. All media were sterilized at 121 ℃ for 20min, cooled and ready for inoculation. Inoculating Pleurotus cornucopiae mycelium on an ultra-clean workbench sterilized by ultraviolet for 30min, culturing at 25 deg.C in dark, culturing with liquid medium in a constant temperature shaking table at 150rpm and 25 deg.C, and culturing with solid medium in a constant temperature incubator at 25 deg.C. The cultivation process of pleurotus cornucopiae is monitored in real time, the growth condition and the colony morphology of the pleurotus cornucopiae are observed, and pictures are taken when hyphae grow to the 5 th day, and the result is shown in the comparative example 1 in the figure 1. In the solid culture process, the growth rate of pleurotus cornucopiae hyphae is evaluated, the radius of a colony is measured by a cross method at 2d and 5d after inoculation, the ratio of the difference (mm) of the radius of the colony to the number (d) of growing days is the growth rate (mm/d) of the hyphae, the three times of the measurement are repeated, and the average measurement result is shown in a comparative example 1 in a table 1. The inoculation amount of the liquid culture medium is 10%, and in the liquid culture process, when the pleurotus cornucopiae mycelium pellets are cultured for 7 days under a dark condition, the pleurotus cornucopiae mycelium pellets basically grow in a triangular flask and are in a vigorous growth stage. The mycelium pellets were filtered through a filter paper dried to a constant weight (M1), the mycelium pellets and the filter paper were dried together to a constant weight (M2), and the dry weight M of the pleurotus cornucopiae mycelium pellets was calculated as (M2-M1) and repeated three times, and the average dry weight of the mycelium pellets was measured as shown in comparative example 1 in table 2. Collecting Pleurotus cornucopiae mycelium in liquid culture medium, determining mycelium laccase activity by ABTS method, extracting protein by lysate method, and performing SDS-PAGE electrophoretic analysis.
Example 1
Example 1 the composition and content of the solid medium were: 200g of potato, 15g of agar, 10g of glucose and 10g of mannitol, distilled water is added to the mixture to be constant volume to 1L, and the composition and content of a liquid culture medium are as follows: 200g of potato, 10g of glucose and 10g of mannitol, adding distilled water to a constant volume of 1L, and using a 250mL triangular flask containing 100mL of liquid culture medium per bottle. All media were sterilized at 121 ℃ for 20min, cooled and ready for inoculation. Inoculating Pleurotus cornucopiae mycelium on an ultra-clean workbench sterilized by ultraviolet for 30min, culturing at 25 deg.C in dark, culturing with liquid medium in a constant temperature shaking table at 150rpm and 25 deg.C, and culturing with solid medium in a constant temperature incubator at 25 deg.C. The cultivation process of Pleurotus cornucopiae was monitored in real time, and the growth and colony morphology of the Pleurotus cornucopiae were observed, and photographed when the hyphae grew to the 5 th day, and the results are shown in example 1 in FIG. 1. In the solid culture process, the growth rate of pleurotus cornucopiae hyphae is evaluated, the radius of a colony is measured by a cross method at 2d and 5d after inoculation, the ratio of the difference (mm) of the radius of the colony to the number (d) of days of growth is the growth rate of the hyphae (mm/d), the three times of the measurement are repeated, and the average measurement result is shown in example 1 in the table 1. The inoculation amount of the liquid culture medium is 10%, and in the liquid culture process, when the pleurotus cornucopiae mycelium pellets are cultured for 7 days under a dark condition, the pleurotus cornucopiae mycelium pellets basically grow in a triangular flask and are in a vigorous growth stage. The mycelium pellets were filtered through a filter paper dried to a constant weight (M1), the mycelium pellets and the filter paper were dried together to a constant weight (M2), and the dry weight M of the pleurotus cornucopiae mycelium pellets was calculated as (M2-M1) and repeated three times, and the average dry weight of the mycelium pellets was measured as shown in example 1 in table 2. Collecting Pleurotus cornucopiae mycelium in liquid culture medium, determining mycelium laccase activity by ABTS method, extracting protein by lysate method, and performing SDS-PAGE electrophoretic analysis.
Example 2
Example 2 the composition and content of the solid medium were: 200g of potato, 15g of agar, 10g of glucose and 20g of mannitol, distilled water is added to the mixture to be constant volume to 1L, and the composition and content of a liquid culture medium are as follows: 200g of potato, 10g of glucose and 20g of mannitol, adding distilled water to a constant volume of 1L, and using a 250mL triangular flask containing 100mL of liquid culture medium per bottle. All media were sterilized at 121 ℃ for 20min, cooled and ready for inoculation. Inoculating Pleurotus cornucopiae mycelium on an ultra-clean workbench sterilized by ultraviolet for 30min, culturing at 25 deg.C in dark, culturing with liquid medium in a constant temperature shaking table at 150rpm and 25 deg.C, and culturing with solid medium in a constant temperature incubator at 25 deg.C. The cultivation process of Pleurotus cornucopiae was monitored in real time, and the growth and colony morphology of the Pleurotus cornucopiae were observed, and photographed when the hyphae grew to the 5 th day, and the results are shown in example 2 in FIG. 1. In the solid culture process, the growth rate of pleurotus cornucopiae hyphae is evaluated, the radius of a colony is measured by a cross method at 2d and 5d after inoculation, the ratio of the difference (mm) of the radius of the colony to the number (d) of days of growth is the growth rate of the hyphae (mm/d), the three times of the measurement are repeated, and the average measurement result is shown in example 2 in the table 1. The inoculation amount of the liquid culture medium is 10%, and in the liquid culture process, when the pleurotus cornucopiae mycelium pellets are cultured for 7 days under a dark condition, the pleurotus cornucopiae mycelium pellets basically grow in a triangular flask and are in a vigorous growth stage. The mycelium pellets were filtered through a filter paper dried to a constant weight (M1), the mycelium pellets and the filter paper were dried together to a constant weight (M2), and the dry weight M of the pleurotus cornucopiae mycelium pellets was calculated as (M2-M1) and repeated three times, and the average dry weight of the mycelium pellets was measured as shown in example 2 in table 2. Collecting Pleurotus cornucopiae mycelium in liquid culture medium, determining mycelium laccase activity by ABTS method, extracting protein by lysate method, and performing SDS-PAGE electrophoretic analysis.
Comparative example 2
Comparative example 2 the composition and content of the solid medium were: 200g of potato, 15g of agar and 50g of mannitol, distilled water is added to the mixture to be constant volume to 1L, and the composition and the content of a liquid culture medium are as follows: 200g of potato and 50g of mannitol, adding distilled water to a constant volume of 1L, and using 250mL triangular flasks, wherein each flask contains 100mL of liquid culture medium. All media were sterilized at 121 ℃ for 20min, cooled and ready for inoculation. Inoculating Pleurotus cornucopiae mycelium on an ultra-clean workbench sterilized by ultraviolet for 30min, culturing at 25 deg.C in dark, culturing with liquid medium in a constant temperature shaking table at 150rpm and 25 deg.C, and culturing with solid medium in a constant temperature incubator at 25 deg.C. The cultivation process of Pleurotus cornucopiae is monitored in real time, the growth and colony morphology are observed, and photographing is carried out when the hypha grows to the 5 th day, and the result is shown in comparative example 2 in FIG. 1. In the solid culture process, the growth rate of pleurotus cornucopiae hyphae is evaluated, the radius of a colony is measured by a cross method at 2d and 5d after inoculation, the ratio of the difference (mm) of the radius of the colony to the number (d) of growing days is the growth rate (mm/d) of the hyphae, the three times of the measurement are repeated, and the average measurement result is shown in a comparative example 2 in a table 1. The inoculation amount of the liquid culture medium is 10%, and in the liquid culture process, when the pleurotus cornucopiae mycelium pellets are cultured for 7 days under a dark condition, the pleurotus cornucopiae mycelium pellets basically grow in a triangular flask and are in a vigorous growth stage. The mycelium pellets were filtered through a filter paper dried to a constant weight (M1), the mycelium pellets and the filter paper were dried together to a constant weight (M2), and the dry weight M of the pleurotus cornucopiae mycelium pellets was calculated as (M2-M1) and repeated three times, and the average dry weight of the mycelium pellets was measured as shown in comparative example 2 in table 2. Collecting Pleurotus cornucopiae mycelium in liquid culture medium, determining mycelium laccase activity by ABTS method, extracting protein by lysate method, and performing SDS-PAGE electrophoretic analysis.
TABLE 1
TABLE 2
FIG. 1 is a graph showing the growth of the hyphae of Pleurotus cornucopiae of examples 1 and 2 and comparative examples 1 and 2 during the culture of Pleurotus cornucopiae. FIG. 1 shows that: the pleurotus cornucopiae mycelia of examples 1 and 2 had better growth vigor than the pleurotus cornucopiae mycelia of comparative example 1, and the pleurotus cornucopiae mycelia of example 2 had the most growing branches and had the best growth vigor, while the pleurotus cornucopiae mycelia of comparative example 2 had sparse mycelia and had similar growth vigor to that of comparative example 1.
FIG. 2 is a graph showing the growth rate of the mycelia of Pleurotus cornucopiae of examples 1 and 2 and comparative examples 1 and 2 during the solid culture of Pleurotus cornucopiae. FIG. 2 shows that: the growth rates of the pleurotus cornucopiae hyphae of the example 1 and the example 2 are very different from those of the comparative example 1, and are both significantly higher than those of the comparative example 2, and the culture medium of the example 2 has the most significant effect of promoting the growth of the pleurotus cornucopiae hyphae.
FIG. 3 is a dry weight chart of the mycelia of Pleurotus cornucopiae of examples 1 and 2 and comparative examples 1 and 2 during the liquid culture of Pleurotus cornucopiae. FIG. 3 shows: the culture media of example 2 and comparative example 2 were able to increase the biomass of pleurotus cornucopiae hyphae, with significant differences between the dry weight of pleurotus cornucopiae hyphae of example 2 and comparative example 2, and no significant difference between the dry weight of pleurotus cornucopiae hyphae of example 1 and comparative example 1.
FIG. 4 is a graph showing laccase activity of Agaricus blazei Murill mycelia of examples 1 and 2 and comparative examples 1 and 2 obtained by liquid culture of Agaricus blazei Murill. FIG. 4 shows that: the laccase activity of the pleurotus cornucopiae mycelia in the examples 1 and 2 is obviously higher than that of the pleurotus cornucopiae mycelia in the comparative example 1, while the laccase activity of the pleurotus cornucopiae mycelia in the comparative example 2 is not obviously different from that of the comparative example 1, and the culture mediums in the examples 1 and 2 can improve the lignin degradation capability of the pleurotus cornucopiae mycelia.
FIG. 5 is a protein electrophoresis chart of Agaricus blazei Murill mycelia in examples 1 and 2 and comparative examples 1 and 2 obtained by liquid culture of Agaricus blazei Murill. FIG. 5 shows that: the pleurotus cornucopiae hypha protein band compositions of examples 1 and 2 were not different from the pleurotus cornucopiae hypha protein band composition of comparative example 1, indicating that the culture media of examples 1 and 2 did not change the protein expression of pleurotus cornucopiae hyphae; the composition of the pleurotus cornucopiae hypha protein band of comparative example 2 is significantly different from that of the pleurotus cornucopiae hypha protein band of comparative example 1, indicating that the culture medium of comparative example 2 changes the protein expression of pleurotus cornucopiae hyphae.
In conclusion, the culture medium in example 2, compared to comparative example 1, while increasing growth rate and hyphal biomass, promoted secretion of hyphal laccase activity and did not affect hyphal protein expression. Example 1 is slightly inferior to example 2 in the hyphal growth state and growth rate; comparative example 2 is significantly inferior to example 2 in the hyphal growth state, growth rate and laccase activity. The above data indicate that the medium formulation in example 2 is the optimal formulation. The formula optimizes the composition of a carbon source in the formula of a basic culture medium (comparative example 1), and the culture effect of the formula with 1% of the addition amount of glucose and 2% of the addition amount of mannitol is superior to that of the formula with 1% of the addition amount of glucose and 1% of the addition amount of mannitol, and is obviously superior to that of the formula with 5% of the addition amount of mannitol.
Claims (7)
1. The pleurotus cornucopiae culture medium comprises a solid culture medium and a liquid culture medium, and is characterized in that the solid culture medium comprises the following components: 200g of potato, 15g of agar, 10g of glucose and 10-20 g of mannitol, and adding distilled water to a constant volume of 1L; the composition of the liquid medium was: 200g of potato, 10g of glucose, 10-20 g of mannitol and distilled water to a constant volume of 1L.
2. The culture medium of Pleurotus cornucopiae according to claim 1, wherein the solid medium and the liquid medium are sterilized at 121 ℃ for 20min and cooled to be inoculated.
3. The culture medium of Pleurotus cornucopiae according to claim 2, wherein the inoculation is carried out on a super clean bench sterilized by UV for 30 min.
4. The culture medium of Pleurotus cornucopiae according to claim 2, wherein the amount of inoculation of the liquid medium is 10%.
5. A method for culturing Pleurotus cornucopiae based on the Pleurotus cornucopiae culture medium according to any one of claims 1 to 4, comprising:
a solid medium or a liquid medium according to any one of claims 1 to 4, wherein the mycelia of Pleurotus cornucopiae are inoculated and cultured in the dark at 25 ℃.
6. The method according to claim 5, wherein the culture is carried out in a shaker at 25 ℃ and a rotation speed of 150rpm while being inoculated into a liquid medium.
7. The culture method according to claim 5, wherein the culture is carried out in an incubator at 25 ℃ while inoculating the culture medium.
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| CN113234605A (en) * | 2021-05-28 | 2021-08-10 | 广东粤微生物科技有限公司 | Simple preparation method of phlebopus portentosus solid and liquid strains |
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