CN113265367B - Culture medium for rapidly obtaining Morchella conidia and preparation method thereof - Google Patents

Abstract

The invention provides a culture medium for rapidly obtaining Morchella conidium and a preparation method thereof. Taking Morchella (Morchella spp.) mother strain obtained by single spore, multiple spores or tissue separation, activating on a common PDA or CYM culture medium to obtain mother strain, and inoculating Morchella into the following formula by a conventional aseptic operation method: culturing in a medium comprising 0-20% of carbon source, 0-20% of nitrogen source, 0.1-2% of trace elements, 12-25% of agar and 5-8% of ph at 5-25deg.C for 14-30 days in dark place, and preserving at 2-8deg.C for use after macroscopic detection or microscopic detection of the culture result.

Description

A kind of cultivation and its preparation method for rapidly obtaining morel conidia

technical field

The invention belongs to the field of biotechnology, and in particular relates to a method for obtaining asexual spores of edible fungus aseptically cultured, including medium ratio and culture conditions. More specifically, it relates to a preparation method for rapidly obtaining morel conidia.

Background technique

Morchella spp. is a world-renowned edible and medicinal fungus, which has the functions of anti-tumor, anti-oxidation, anti-bacterial, lowering blood fat, and improving immunity. In recent years, with people's pursuit of green health, morels have been praised by more and more people, and the demand is increasing day by day. The number of wild morels has decreased sharply, and artificial cultivation of morels has increasingly become the focus of research. In 2010, the popularization and dissemination of independent nutrient addition technology made morel cultivation technology develop rapidly. However, nearly 70% of growers still cannot obtain stable income every year. The main reason is that morel species, life history, nutrient metabolism, physiology and other basic biological research is lacking.

Conidia is the most common type of asexual spore among fungi with septal hyphae. It is the asexual reproduction mode of most Ascomycotina and all Deuteromycotina fungi. It is mainly transmitted by wind, water and animals. . Characteristics such as spore morphology, structure, size, color and arrangement vary from species to species, and are an important basis for strain identification. Some conidia can be used for breeding, preservation and inoculation expansion.

The earliest report of morel conidia was in the artificial cultivation experiment attempted by Molliard in 1904. The experiment found that a large amount of white mold could be formed on the surface of the inoculated soil layer. In 1982, a large number of conidia were also reported in Ower's morel artificial cultivation experiment, and pointed out that its germination rate on potato medium was less than 1%. In 2005, Masaphy first described the asexual spore morphology of Morchella rufica during cultivation. The commercial cultivation of Morchella shows that the production of conidia (commonly referred to as "bacteria cream") has an important indicator effect on fruiting. Morchella is very easy to produce conidia during field cultivation (Figure 4). However, except for a small amount of conidia obtained by Qi Peng et al. (2021) using sterilized humus soil medium, there is no mother species of Morchella Reports on obtaining conidia stably and rapidly under other sterile conditions.

Contents of the invention

The object of the present invention is to provide a kind of pure culture method of hickory chick conidia aiming at the above-mentioned deficiencies existing in the prior art. The method of the invention can obtain conidia of hickory chick under aseptic conditions, solves the problem of difficulty in obtaining conidia of hickory chick through pure culture, and provides a standard for selection and breeding of hickory chick good varieties. At the same time, it provides a culture medium for quickly obtaining conidia of Morchella, breaks through the limitations of difficult and easy-to-contaminate pure culture of Morchella, and provides a scientific method for the selection and breeding of Morchella varieties.

Above-mentioned purpose of the present invention is achieved by following technical scheme:

A preparation method for rapidly obtaining conidia of hickory chick, the method comprises the steps of preparation of special medium, inoculation, cultivation and detection, etc., taking single spore, polyspore or tissue isolation and obtaining hickory chick mother seed in common PDA or Activated on the CYM medium as the mother species, and then routinely inoculated into the formula: 0-20% carbon source, 0-20% nitrogen source, 0.1-2% trace elements, agar 12-25%, ph5-8 On the culture medium, culture at 5-25°C in the dark for 14-30 days, and store at 2-8°C for later use after visual inspection or microscopic inspection of the culture results.

According to the preparation method for quickly obtaining morel conidia, the preparation method of the special medium is: 0-20% carbon source, 0-10% nitrogen source, 0.5-2% trace elements, agar 12-25%, 1000ml of water, pH 5-8, sterilized at 121-126°C for 15-90 minutes, and then made into a culture medium by conventional parent seed production methods.

According to the preparation method for rapidly obtaining morel conidia, the culturing step is culturing at 5-25 degrees for 14-30 days after inoculation until the hyphae are completely covered with petri dishes or test tubes.

According to the preparation method for rapidly obtaining morel conidia, the carbon source is one or a mixture of at least two of wheat, sawdust, cottonseed hulls, glucose, sucrose and fructose.

According to the preparation method for rapidly obtaining morel conidia, the nitrogen source is one or a mixture of at least two of yeast powder, peptone, beef extract and soybean powder.

According to a kind of preparation method of obtaining morel conidia rapidly, wherein said trace element is one or at least two mixtures of magnesium sulfate, potassium dihydrogen phosphate, dipotassium hydrogen phosphate and potassium carbonate .

The invention also provides a special culture medium for quickly obtaining morel conidia, the formula of the special culture medium is: 0-20% carbon source, 0-10% nitrogen source, 0.5-2% trace elements, Agar 12-25%, water 1000ml, pH5-8.

A special medium for quickly obtaining morel conidia, wherein the special medium is prepared by the following formula and method: 0-20% carbon source, 0-10% nitrogen source, 0.5-2% trace Elements, agar 12-25%, water 1000ml, pH 5-8, sterilized at 121-126°C for 15-90min, and the conventional mother seed production method was used to prepare the culture medium.

The invention provides a rapid, low-cost, simple procedure, high repeatability, and easy-to-observe method for pure cultivation of hickory chick conidia, and simultaneously provides a culture medium for quickly obtaining hickory chick conidia. The method of the present invention can obtain morel conidia under aseptic conditions, solves the technical problem in the prior art that it is difficult to obtain conidia by pure culture of morel, and provides a breeding standard for morel varieties . The invention breaks through the limitations of the difficulty in pure culture of the hickory chick and the easy contamination of the cultivation, and provides a scientific and operable method for the selection and breeding of the hickory chick.

Description of drawings

Fig. 1 is No. 13 bacterial strain conidia of Liumei hickory chick;

Fig. 2 is the conidia of the 301 bacterial strains of the morerella edulis;

Fig. 3 is the conidia of Qimei hickory chick 202 bacterial strain;

Figure 4 shows morel conidia (commonly referred to as "bacteria cream").

Detailed ways

Below in conjunction with the accompanying drawings, the present invention will be further described with the embodiments of the present invention, but the embodiments are only for illustration and cannot limit the scope of the present invention.

Example 1:

A culture medium for rapidly obtaining morel conidia and its preparation method according to the present invention, comprising: on June 10, 2020, the common PDA culture medium was used as the material of Liumei Morchella No. 13 strain which is common in the market After activation, it became the mother species. On June 16, it was transferred to a medium containing 20% glucose, 5% yeast powder, 0.1% potassium carbonate, 0.3% magnesium sulfate, 20% agar, and pH7, and cultured at 20°C in the dark for 13 days. days, conidia as shown in Figure 1 can be obtained.

Example 2:

A culture medium for quickly obtaining morel conidia and its preparation method according to the present invention, comprising: on April 10, 2020, the common CYM culture medium was used to activate the common CYM medium The latter is the mother species, and it was transferred on April 15 to a medium containing 10% sucrose, 10% glucose, 2% yeast powder, 3% peptone, 0.2% potassium carbonate, 0.3% magnesium sulfate, 18% agar, and pH7 , and cultured at 15°C in the dark for 17 days, conidia as shown in Figure 2 can be obtained.

Example 3:

A culture medium for rapidly obtaining morel conidia and its preparation method according to the present invention, comprising: on October 9, 2020, the common PDA culture medium was used to activate the common PDA culture medium, taking the seven-mei morel 202 strain as the material The latter is the mother species. On October 14, it was transferred to a medium containing 8% wheat, 2% yeast powder, 0.1% potassium dihydrogen phosphate, 0.1% magnesium sulfate, 18% agar, pH 7, and cultured at 25°C in the dark After 14 days, conidia as shown in Figure 3 can be obtained.

Claims (3)

1. The preparation method comprises the steps of preparing a special culture medium, inoculating, culturing and detecting, wherein Morchella strains obtained by single spore, multiple spores or tissue separation are activated on a common PDA or CYM culture medium to form mother seeds, and then the mother seeds are inoculated to a formula by conventional aseptic operation: 20% glucose, 5% yeast powder, 0.1% potassium carbonate, 0.3% magnesium sulfate, 20% agar, 1000ml water and pH7, culturing in dark at 15-25deg.C for 14-30 days, and preserving at 2-8deg.C for use after macroscopic or microscopic detection of the culture result.

2. The preparation method comprises the steps of preparing a special culture medium, inoculating, culturing and detecting, wherein Morchella strains obtained by single spore, multiple spores or tissue separation are activated on a common PDA or CYM culture medium to form mother seeds, and then the mother seeds are inoculated to a formula by conventional aseptic operation: 10% of sucrose, 10% of glucose, 2% of yeast powder, 3% of peptone, 0.2% of potassium carbonate, 0.3% of magnesium sulfate, 18% of agar, 1000ml of water and pH7 are cultured for 14-30 days in a dark place at 15-25 ℃, and after visual inspection or microscopic inspection of the culture result, the culture is preserved at 2-8 ℃ for later use.

3. The preparation method comprises the steps of preparing a special culture medium, inoculating, culturing and detecting, wherein Morchella strains obtained by single spore, multiple spores or tissue separation are activated on a common PDA or CYM culture medium to form mother seeds, and then the mother seeds are inoculated to a formula by conventional aseptic operation: 8% of wheat, 2% of yeast powder, 0.1% of monopotassium phosphate, 0.1% of magnesium sulfate, 18% of agar, 1000ml of water and pH7 are cultured for 14-30 days at 15-25 ℃ in a dark place, and after the culture result is detected by naked eyes or microscopy, the culture is preserved at 2-8 ℃ for later use.

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