CN111454850A - Simple preservation method for edible and medicinal fungi strains - Google Patents
Simple preservation method for edible and medicinal fungi strains Download PDFInfo
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- CN111454850A CN111454850A CN202010446499.4A CN202010446499A CN111454850A CN 111454850 A CN111454850 A CN 111454850A CN 202010446499 A CN202010446499 A CN 202010446499A CN 111454850 A CN111454850 A CN 111454850A
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- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
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- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/04—Preserving or maintaining viable microorganisms
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Abstract
The invention relates to the preservation of edible fungus strains, in particular to a simple and convenient preservation method of edible and medicinal fungus strains, which comprises the following steps: (1) inoculating the strain into a plate culture medium, uniformly punching the cultured plate strain by using a sterile puncher to prepare a strain block with the diameter of 0.4-0.5 cm, and uniformly inoculating the strain block into a new plate culture medium; (2) flatly paving filter paper sheets around the bacterium blocks, and keeping a distance between the filter paper sheets; (3) culturing the inoculated plate until the hyphae completely cover the filter paper sheet; (4) slightly lifting the filter paper sheet covered with the edible and medicinal fungi hyphae without adhering culture medium, and transferring the filter paper sheet to an aseptic kraft paper envelope; (5) and putting the kraft paper envelope filled with the hypha filter paper sheets into allochroic silica gel for drying, placing the kraft paper envelope in a closed container at normal temperature for 10-15 days, sealing the kraft paper envelope, and transferring the kraft paper envelope to a 4 ℃ short-term storage or express distribution. The method of the invention is simple and easy to operate, has good hypha consistency, is used for short-term preservation, and is beneficial to follow-up research.
Description
Technical Field
The invention relates to preservation of edible fungus strains, in particular to a simple and convenient preservation method of edible and medicinal fungus strains.
Background
The edible and medicinal fungi strains are important basic stones for the development of the edible and medicinal fungi industry, and the quality of the strains directly influences the yield and the quality of the edible and medicinal fungi sporocarp. At present, the preservation method of edible and medicinal fungi strains mainly comprises subculture cryopreservation and ultralow temperature (liquid nitrogen) preservation, and most of media used mainly comprise glass test tubes (slant culture medium) and freezing tubes (liquid culture medium containing cell protection solution). These preservation methods are certainly advantageous for long-term preservation of the strains, but are extremely inconvenient in production and circulation, and require special protection devices or containers for transportation.
Disclosure of Invention
Aiming at the defects, the invention provides a simple and convenient short-term strain preservation method, which is beneficial to the transportation and circulation of strains in production.
The invention achieves the above purposes through the following scheme:
a simple preservation method for edible and medicinal fungi strains comprises the following steps:
(1) inoculating the strain into a plate culture medium, uniformly punching the cultured plate strain by using a sterile puncher to prepare a strain block with the diameter of 0.4-0.5 cm, and uniformly inoculating the strain block into a new plate culture medium;
(2) flatly paving filter paper sheets around the bacterium blocks, and keeping a distance between the filter paper sheets;
(3) culturing the inoculated plate until the hyphae completely cover the filter paper sheet;
(4) slightly lifting the filter paper sheet covered with the edible and medicinal fungi hyphae without adhering culture medium, and transferring the filter paper sheet to an aseptic kraft paper envelope;
(5) and putting the kraft paper envelope filled with the hypha filter paper sheets into allochroic silica gel for drying, placing the kraft paper envelope in a closed container at normal temperature for 10-15 days, sealing the kraft paper envelope, and transferring the kraft paper envelope to a 4 ℃ short-term storage or express distribution.
The strain preserved by the method can be preserved for 1-2 months at 4 ℃ in a short period or can be preserved for 1-2 months at normal temperature, so that the strain can be conveniently used for express distribution and transportation.
Preferably, the filter paper sheet is medium-speed quantitative filter paper, so that mycelia can be attached conveniently.
Preferably, the filter paper sheet is a square or round filter paper sheet.
In one embodiment, the filter paper sheet is a 1cm by 1cm or 2cm by 2cm square filter paper sheet.
In one embodiment, the filter paper sheet is a circular filter paper sheet of 1cm by 1cm or 2cm by 2 cm.
Preferably, the strain is inoculated into a plate culture medium with a diameter of 9 cm.
Preferably, the culture medium is a PDA culture medium.
Preferably, the filter paper sheets are flatly laid around the bacterium blocks, and a distance of 0.5-1 cm is reserved between the filter paper sheets.
Preferably, the envelope is an aseptic self-made filmed kraft envelope.
In the preservation method provided by the invention, the strain blocks are uniformly inoculated to the new plate culture medium, so that the mycelium can quickly grow on the whole plate as far as possible and uniformly cover the whole plate to the filter paper sheet, the defect of inconsistent growth period of the mycelium caused by different positions of the filter paper sheet is avoided, the strain quality is influenced, the parent strains or primary strains with the same source are propagated or the strains are reserved in the mode, the mycelium is ensured to grow fresh and synchronously as far as possible, and the variation of the strain caused by different growth periods is avoided to the greatest extent, and the stability of the agronomic characters in production is influenced.
In the preservation method provided by the invention, the filter paper sheets full of hyphae are dried for 1-2 weeks by using allochroic silica gel, so that the hyphae are properly dried, the metabolic rate of the hyphae is reduced, and the ageing of the hyphae is delayed.
In the preservation method provided by the invention, the inoculated strains are kept in non-contact with the filter paper, so that the hyphae spreading to the filter paper sheet are fresh hyphae growing out.
The invention adopts multi-point uniform inoculation, mycelium grows over the flat plate quickly and uniformly, and the aging or variation of hyphae grown first caused by over-long growth time is avoided, thereby ensuring the consistency of the preserved hyphae, furthermore, the filter paper sheet is not in contact with the inoculated strains, the adhesion of the inoculated hyphae on the filter paper sheet is avoided, the hyphae on the filter paper sheet are ensured to grow out fresh, and the consistency of the hyphae is maintained.
The method of the invention is simple and easy to operate, has good hypha consistency, is used for short-term preservation, and is beneficial to subsequent research.
Drawings
FIG. 1 is a schematic representation of the seed patch inoculation site.
Figure 2 is one of the schematic representations of the homemade process of a sterile laminated (PE) kraft envelope.
Figure 3 is a second illustration of the homemade process of sterile film-coated (PE) kraft envelopes.
Figure 4 is a third illustration of the homemade process of a sterile film-coated (PE) kraft envelope.
Detailed Description
The present invention is further illustrated by the following specific examples.
Example 1:
1. preparing a square or round filter paper sheet (limited to medium-speed quantitative filter paper for facilitating adhesion of mycelia) with the specification of 1cm x 1cm or 2cm x 2cm, using sterile tweezers, self-making a sterile film-coated (PE) kraft paper envelope, and placing the envelope on an ultra-clean workbench for later use;
2. preparing a nutrient medium required by edible and medicinal fungi strains, sterilizing at high temperature and high pressure (such as 121 ℃, 20min), packaging into a dish with a diameter of 9cm in a super clean bench, and cooling and solidifying for later use;
3. the cultured flat strain is uniformly punched by a 0.5cm sterile puncher to prepare a strain block with the diameter of 0.5cm, the strain block is uniformly inoculated into a new PDA flat culture medium according to the marked position shown in figure 1, then sterile filter paper sheets are flatly laid around the strain block by using sterile tweezers, a proper distance (about 0.5cm) is reserved between the filter paper sheets, and the strain block is inoculated according to the position shown in figure 1, so that the problem that the mycelium grows over the whole flat plate as quickly as possible and covers the filter paper sheets uniformly is solved, and the defect that the growth period of the mycelium is inconsistent due to different positions of the filter paper sheets is avoided, and the strain quality is influenced. Keeping the inoculated strains and the filter paper in a non-contact manner so as to ensure that hyphae spreading to the filter paper sheet are grown fresh hyphae;
4. sealing the inoculated plate with sealing film (or sealing adhesive tape), culturing in artificial climate incubator in dark place until the mycelium completely covers the filter paper sheet, and adjusting the culture temperature according to the characteristics of edible and medicinal fungi;
5. preparing sterile tweezers and a sterile waterproof kraft paper bag (about 7 x 9cm), and placing the bag on an ultra-clean workbench for later use;
6. in a clean bench, gently lifting the filter paper sheet covered with edible and medicinal fungi hyphae with sterile tweezers (taking care not to stick culture medium), transferring to a sterile self-made laminating (PE) kraft paper envelope, and folding the envelope for 2-3 times;
7. putting kraft paper envelopes containing hypha filter paper sheets into allochroic silica gel for drying, placing the envelopes in a closed container at normal temperature for about 2 weeks, then moderately drying the hypha), pasting sealing adhesive tapes on the paper bag roll mouths, and moving the envelopes into enterprises or producers needing strains for short-term storage at 4 ℃ for 1-2 months or express transportation circulation (for 1-2 months)
The method comprises the following steps of (1) self-making aseptic coated (PE) kraft paper envelope process:
① cutting off the kraft paper with a film (PE) as shown in FIG. 2;
② As shown in FIG. 3, regions ①, ② and ③ are folded in sequence along the dashed lines in the direction of the arrows to form side creases in the intermediate region;
③ transferring the kraft paper to a clean bench, sterilizing with surface alcohol, standing, irradiating with ultraviolet rays on both sides for 30 min, and air drying;
④ As shown in FIG. 4, the overlapping portions (shaded) of regions ①, ②, ③ are bonded together at a clean bench using sterile double-sided tape and stored in a sterile condition until completely bonded together, and region ④ is the flap portion for sealing during use.
Example 2: use of strains with filter paper as preservation medium
1. Preparing sterile tweezers, sterile scissors and a sterile culture medium (a plate or a test tube);
2. sterilizing the surface of the small paper bag filled with the strain by using 75% alcohol, transferring the small paper bag to a super-clean workbench, cutting the opening of the bag by using sterile scissors, taking out the filter paper sheet covered with hyphae by using sterile tweezers, and connecting the filter paper sheet to a plate or a test tube culture medium;
3. and transferring the inoculated plate or test tube culture medium to an artificial climate incubator, and performing dark culture at a proper temperature until hyphae grow over the plate or test tube, and taking out for use.
The above description is only for the preferred embodiment of the present invention, but the scope of the present invention is not limited thereto, and any person skilled in the art should be considered to be within the technical scope of the present invention, and the technical solutions and their concepts should be considered to be equivalent or modified within the technical scope of the present invention.
Claims (9)
1. A simple preservation method for edible and medicinal fungi strains is characterized by comprising the following steps:
(1) inoculating the strain into a plate culture medium, uniformly punching the cultured plate strain by using a sterile puncher to prepare a strain block with the diameter of 0.4-0.5 cm, and uniformly inoculating the strain block into a new plate culture medium;
(2) flatly paving filter paper sheets around the bacterium blocks, and keeping a distance between the filter paper sheets;
(3) culturing the inoculated plate until the hyphae completely cover the filter paper sheet;
(4) slightly lifting the filter paper sheet covered with the edible and medicinal fungi hyphae without adhering culture medium, and transferring the filter paper sheet to an aseptic kraft paper envelope;
(5) and putting the kraft paper envelope filled with the hypha filter paper sheets into allochroic silica gel for drying, placing the kraft paper envelope in a closed container at normal temperature for 10-15 days, sealing the kraft paper envelope, and transferring the kraft paper envelope to a 4 ℃ short-term storage or express distribution.
2. The convenient preservation method for edible and medicinal fungi strains according to claim 1, characterized in that the filter paper sheet is medium-speed quantitative filter paper.
3. The method for conveniently preserving edible and medicinal fungi strains according to claim 1, wherein the filter paper sheet is a square or round filter paper sheet.
4. The method of claim 3, wherein the filter paper is a 1cm by 1cm or 2cm by 2cm square filter paper.
5. The method of claim 4, wherein the filter paper sheet is a circular filter paper sheet of 1cm by 1cm or 2cm by 2 cm.
6. The method for the convenient preservation of edible and medicinal fungi strains according to claim 1, wherein the strains are inoculated into a plate culture medium with a diameter of 9 cm.
7. The method for conveniently preserving edible and medicinal fungi strains according to claim 1, wherein the culture medium is a PDA culture medium.
8. The convenient edible and medicinal fungi strain preservation method according to claim 1, wherein the filter paper sheets are laid around the fungi block, and a distance of 0.5-1 cm is reserved between the filter paper sheets.
9. The convenient preservation method for edible and medicinal fungi strains according to claim 1, characterized in that the envelope is a sterile self-made coated kraft paper envelope.
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| Application Number | Priority Date | Filing Date | Title |
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| CN202010446499.4A CN111454850A (en) | 2020-05-25 | 2020-05-25 | Simple preservation method for edible and medicinal fungi strains |
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| Application Number | Priority Date | Filing Date | Title |
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| CN202010446499.4A CN111454850A (en) | 2020-05-25 | 2020-05-25 | Simple preservation method for edible and medicinal fungi strains |
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| CN111454850A true CN111454850A (en) | 2020-07-28 |
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN112899166A (en) * | 2021-04-06 | 2021-06-04 | 华中农业大学 | Fungus strain preservation method |
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2020
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| CN109207375A (en) * | 2018-10-09 | 2019-01-15 | 山西大学 | A kind of method for preserving of plant endogenesis epiphyte strain |
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN112899166A (en) * | 2021-04-06 | 2021-06-04 | 华中农业大学 | Fungus strain preservation method |
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Application publication date: 20200728 |