A sanguinea var. iris strain Qian P216 and its application
By optimizing the culture method and cultivation conditions of the winter iris strain Guizhou P216, the problems of long winter iris cultivation cycle and low yield were solved, and efficient and stable winter iris production was achieved.
Patent Information
- Application Number
- CN202510094973.4
- Authority / Receiving Office
- CN · China
- Patent Type
- Patents(China)
- Current Assignee / Owner
- Filing Date
- 2025-01-21
- Publication Date
- 2025-09-19
- Estimated Expiration
- 2045-01-21
AI Technical Summary
There are few existing winter iris cultivars, the mycelium grows slowly, the production cycle is long, and the traits are unstable, resulting in unstable yield and quality, which cannot meet market demand and has high costs.
Provided is a winter iris strain Guizhou P216, which optimizes cultivation conditions, shortens cultivation cycles and increases yields through cultivation methods of mother species, original species and cultivars, combined with an understory cultivation model.
The cultivation period of strain Guizhou P216 was shortened to 7-8 months, and the yield was increased to 2.33kg/m2, which was significantly higher than that of existing strains. The nutritional value was improved, labor costs were reduced, and the impact on the climate and environment was reduced.
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Figure CN119899751B_ABST
Abstract
Description
Technical Field
[0001] The invention belongs to the field of edible fungi, and in particular relates to a winter iris strain and also relates to the use of the strain. Background Art
[0002] Dongsun (Phallus dongsun, also known as white ghost pen, bamboo mushroom, bamboo mushroom, and skirtless sunflower) belongs to the genus Phallus in the family Phallaceae and is a well-known medicinal and edible fungus in my country. Dongsun is primarily found in Guizhou, Sichuan, Yunnan, Anhui, and Guangdong. Dongsun has a crisp texture, a delicious flavor, and is rich in nutrients, including a variety of amino acids, proteins, vitamins, and polysaccharides. Dongsun is also a traditional folk medicine in Asia, known for its blood circulation, pain relief, antibacterial properties, edema reduction, and the treatment of rheumatic joint pain. Its active ingredients also have immunomodulatory and wound healing properties. With improvements in living standards and a growing awareness of the nutritional and medicinal value of Dongsun, demand for Dongsun has increased significantly, but current production is insufficient to meet demand. Currently, the market price of Dongsun is 280-450 yuan per kilogram, far higher than the price of other edible fungi. Dongsun has become a key source of income for local farmers.
[0003] The main problems with winter iris cultivation in my country are as follows: (1) There are few cultivated varieties. (2) The mycelium grows slowly, the production cycle is long, and the traits are unstable. Currently, the production cycle of the main cultivated winter iris strains is generally as long as 9 to 12 months, which requires a lot of labor costs, increases the impact of climate and environment, and increases the risk of pests and diseases, resulting in unstable winter iris yield and quality. Therefore, in order to meet market demand and reduce costs, breeding new winter iris strains with short cultivation cycles and high yields is the best way to solve the problems in winter iris production.
[0004] The pristine forests of southern my country, including Guizhou, Sichuan, and Yunnan, are rich in wild edible mushrooms, including wild winter clover. The development and domestication of wild winter clover strains not only enriches people's dining tables and meets their needs for a higher standard of living, but also leverages their medicinal and edible properties to develop higher-value-added foods, beverages, health products, and medicines. This, while improving people's health and well-being, can also increase mushroom farmers' incomes and export earnings. Summary of the Invention
[0005] The invention aims to provide a winter iris strain with short cultivation period and high yield.
[0006] Another object of the present invention is to provide uses of the aforementioned Ilex strychnifolia strain.
[0007] The purpose of the present invention is achieved through the following technical solutions:
[0008] The invention provides a Phallus dongsun strain Qian P216, which is preserved in the General Microbiology Center of China Culture Collection of Microorganisms, and its preservation number is CGMCC No.40102.
[0009] The present invention also provides the use of the strain Guizhou P216 in food or health products.
[0010] The present invention also provides a food or health product, which contains the strain Guizhou P216 or an extract of Guizhou P216.
[0011] The present invention also provides application of the strain Guizhou P216 in beverages.
[0012] The present invention also provides a beverage containing the above-mentioned strain Guizhou P216 or an extract of Guizhou P216.
[0013] The present invention also provides the use of the strain Guizhou P216 in preparing medicines for improving human immunity.
[0014] The present invention also provides the use of the strain Guizhou P216 in preparing medicines for promoting blood circulation, relieving pain, inhibiting bacteria, eliminating edema or treating rheumatic joint pain.
[0015] The present invention also provides a genetically engineered winter iris strain, the starting strain of which is the above-mentioned strain Qian P216.
[0016] The present invention also provides a cultivation method of the above-mentioned winter iris strain Guizhou P216, comprising the following steps:
[0017] (1) Stock culture: The stock of the strain Guizhou P216 was transferred to the stock culture medium, and then placed in a 25°C constant temperature incubator for dark culture for 20 to 25 days, until the mycelium covered the culture medium, and the Guizhou P216 stock was obtained; the components of the stock culture medium were: 200 g potato, 20 g glucose, 20 g agar, 0.3 g potassium dihydrogen phosphate, 0.15 g magnesium sulfate, 10 mg vitamin B1, and 1000 mL water; after preparation, the mixture was sterilized at 121°C for 30 min and poured into a 9 cm diameter plate;
[0018] (2) Stock culture: The culture medium plate covered with the mother strain of Guizhou P216 obtained in step (1) is streaked with bacterial blocks using the well-shaped method and transferred to the stock culture medium, with one plate transferred to one bottle, and then cultured at 25°C in the dark for about 50 days until the mycelium fills the strain bottle, thereby obtaining the Guizhou P216 stock; wherein the components of the stock culture medium are: 89% miscellaneous sawdust (fine), 10% wheat bran, 1% lime; and 70% water content; during preparation, the components are mixed uniformly according to the proportion, and then water is added according to the proportion; finally, the mixture is placed in a 500mL glass strain bottle, and sterilized under high pressure at 121°C for 1h. After sterilization, the mixture is placed in a clean bench and cooled to room temperature;
[0019] (3) Cultivation of cultivars: The original Guizhou P216 obtained in step (2) is transferred to a culture medium, with a transfer of about 15 g per bag, and cultured at 25°C in the dark for about 50 days until the fungus bag is full, thereby obtaining the Guizhou P216 cultivar; wherein the composition of the culture medium is: 80% miscellaneous sawdust, 18% wheat bran, 1% lime, and 1% gypsum; and the water content is 65%; during preparation, the components are mixed evenly according to the proportion, and then water is added according to the proportion; the prepared culture medium is placed in a polypropylene plastic bag (16 cm × 35 cm × 0.005 cm), tied with a polypropylene rope, and sterilized under high pressure at 121°C for 2 h, and cooled to room temperature after the sterilization is completed; wherein the miscellaneous sawdust is coarse sawdust and needs to be pre-wetted in advance to prevent contamination caused by incomplete sterilization;
[0020] (4) Land preparation and bed construction: The cultivation of Guizhou P216 is carried out in an under-forest cultivation mode. The cultivation site is selected in a forest with a high density. The soil should be fertile, loose, and have a relative humidity of 70% to 85%. It is best to have a certain slope to facilitate drainage. When digging furrows, the width should be 50 cm, the depth should be 15 to 20 cm, and the length should be unlimited. The bottom of the furrow should be leveled, and 3 to 4 cm thick loose soil should be laid to retain water. The width between the two furrows should be 20 to 25 cm.
[0021] (5) Paving material: The cultivation method of Guizhou P216 is to use a layer of wood, a layer of fungus, and then a layer of wood. The wood is fresh broad-leaved trees, such as chestnut trees and cycads. They are cut into blocks 2 to 5 cm thick and 5 to 10 cm long. The wood is then evenly spread on the bottom of the ditch and compacted to cover the soil in the ditch.
[0022] (6) Sowing: The sowing time is from March to May. Break the Guizhou P216 cultivar obtained in step (3) into small pieces with a diameter of 3 to 5 cm and sow them on the bottom wood block with a spacing of 3 to 4 cm between each cultivar. Then cover them with a thin layer of bamboo leaves and then a layer of wood.
[0023] (7) Covering soil: Choose clean, loose soil without obvious bacterial growth. The thickness of the covering soil should be 3 to 5 cm. In drought-prone areas, the covering soil should be slightly thicker, 5 to 8 cm. The covering soil should be in the shape of a turtle's back. Then, add a layer of pine needles about 2 cm thick to moisturize the soil.
[0024] (8) Management of mushroom production: The optimal temperature for the growth of Guizhou P216 mycelium is 16-24°C, and the temperature for the formation of Guizhou P216 fruiting bodies is generally 5-12°C. Guizhou P216 has different requirements for humidity at different growth stages. During the mycelium growth stage, the humidity is 70%-85%, the relative humidity is above 80% during the bud differentiation stage, and the soil humidity is 80%-85% during the fruiting body growth stage, and the relative humidity is above 80%. Winter fungus does not require much light for growth, and the cultivation environment requires a semi-shaded and semi-sunny environment for best results. Avoid direct sunlight that may damage the mushroom buds.
[0025] (9) Harvesting: The fruiting body of Guizhou P216 is formed from October to December, and the entire growth cycle from sowing to harvesting is 7 to 8 months. Winter fungus is fragile and easy to break, so be especially careful when harvesting. First, cut off the bottom mycelium with a knife, then gently turn and pull it up. Do not pull it directly to avoid damaging the mycelium and affecting the secondary fruiting.
[0026] The advantages and beneficial technical effects of the present invention are as follows: (1) The stipes of the winter clover strain Guizhou P216 of the present invention are white, cylindrical and hollow, with good mushroom shape and high commercial value. (2) The mushrooms are large. The stipes of Guizhou P216 are 20 to 28 cm long and 3 to 6 cm in diameter. The average weight of fresh mushrooms is 58.27 g / piece, which is significantly higher than the fresh mushroom weight of 25 to 40 g / piece of other existing winter clover strains. (3) The cultivation cycle is short. The cultivation cycle of the strain Guizhou P216 of the present invention from sowing to the beginning of mushroom production and harvesting is 7 to 8 months, while the cultivation cycle of other existing winter clover strains is generally 9 to 12 months. The cultivation cycle of the strain of the present invention is 2 to 4 months shorter than that of other existing winter clover strains; it not only saves labor costs, but also reduces the impact of climate and environment and the harm of pests and diseases, ensuring the stability of fresh mushroom yield and quality. (4) High yield. The fresh product yield of other existing winter clover strains is generally 1.0 to 1.5 kg / m 2 The average fresh product yield of the strain Guizhou P216 of the present invention is 2.33 kg / m 2 , which is significantly higher than the fresh product yield of other existing winter iris strains. (5) High nutritional value. The total sugar content of the fresh mushrooms of the strain Qian P216 of the present invention is 28.81%, which is significantly higher than the total sugar content of other existing winter iris strains of 5-20%; the crude protein content of Qian P216 is 17.86%, which is higher than the crude protein content of other existing winter iris strains of 8-16%.
[0027] Biological preservation: The Phallus dongsun strain Guizhou P216 of the present invention is a wild strain collected by the inventor in Dafang County, Bijie City, Guizhou Province in 2018, and was artificially domesticated and cultivated; Guizhou P216 was deposited in the General Microbiology Center of the China Culture Collection Administration on February 17, 2022, at the Institute of Microbiology, Chinese Academy of Sciences, No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, and its deposit number is CGMCC NO.40102. BRIEF DESCRIPTION OF THE DRAWINGS
[0028] Figure 1 .Photo of the fruiting body of the winter iris strain Guizhou P216 of the present invention.
[0029] Figure 2 . Phylogenetic tree diagram of the winter iris strain Guizhou P216 of the present invention. DETAILED DESCRIPTION
[0030] The present invention is further described below by way of examples, which do not constitute any limitation to the present invention. Unless otherwise specified, the chemical reagents used in the following examples can be purchased from the market.
[0031] Example 1 Collection, Isolation and Domestication Process of the Strain Guizhou P216 of the Present Invention
[0032] In October 2018, the inventors collected multiple wild winter sun fruiting bodies in the wild in Yangchang Town, Dafang County, Bijie City, Guizhou Province, and obtained mycelium (species) by fruiting body tissue separation. After morphological characteristics, ITS-PCR detection, phylogenetic tree construction, and gene fragment size and sequence, it was identified as winter sun (Phallus dongsun). The strains obtained by tissue separation were cultivated from 2019 to 2021, and the fruiting bodies with good commercial traits were tissue separated every year. They were all cultured on the mother culture medium (the mother culture medium formula is: 200g potato, 20g glucose, 20g agar, 0.3g potassium dihydrogen phosphate, 0.15g magnesium sulfate, 10mg vitamin B1, 1000mL water. After preparation, it was autoclaved at 121°C for 30min and poured into a 9cm diameter plate) to screen out strains with strong mycelial growth, fast growth rate and uniform colony, and then used as mother cultures for the next year's cultivation. In 2021, a strain with a well-developed mushroom shape, high yield, and a short production cycle was selected and named Guizhou P216. The fruiting body of this strain has a cap 2 to 7 cm tall and 2 to 7 cm in diameter. The cap has a lattice-like surface, within which dark green, sticky sporophytes are attached. The spores are oval to oblong, smooth, and colorless. The stem is white, spongy, hollow, 20 to 28 cm long, and 3 to 6 cm in diameter. The trophicula is membranous, off-white, 4 to 8 cm tall, and 5 to 12 cm in diameter.
[0033] Example 2 Classification and Identification of the Strain Guizhou P216 of the Present Invention
[0034] (1) Morphological identification of the strain Guizhou P216 of the present invention
[0035] The present strain, Guizhou P216, has solitary or clustered buds, a sterile fruiting body with a bell-shaped cap that becomes flat and perforated after maturity. The cap has a lattice-like surface, with dark green, sticky sporophytes attached within the lattice. The spores are smooth, oval to oblong, and colorless and transparent. The stipe is white, spongy, hollow, and has a lattice-like center. The pilus is off-white and has cords. According to the description and photos of the morphological characteristics of Phallus dongsun on page 478 of "Chinese Edible Fungi Cultivation" (Zhang Jinxia et al., China Agriculture Press, 2020), the present strain, Guizhou P216, has the same morphological characteristics as Phallus dongsun, indicating that the present strain, Guizhou P216, belongs to the genus Phallus dongsun.
[0036] (2) Molecular biological classification and identification of the strain Guizhou P216 of the present invention
[0037] The genomic DNA of the Guizhou P216 strain was extracted using the new plant genomic DNA extraction kit CW0531 (Beijing Kangwei Century Company). ITS-PCR amplification was performed using universal primers ITS4 and ITS5 to obtain the amplified product. The primer sequences for ITS-PCR amplification are as follows:
[0038] ITS5: 5'-GGAAGTAAAAGTCGTAACAAGG-3';
[0039] ITS4: 5'-TCCTCCGCTTATTGATATGC-3'.
[0040] The ITS-PCR amplification system (20 μL) is as follows: 1 μL of 20-50 ng / μL DNA template, 10× Buffer (with Mg 2+ ) 2 μL, 0.5 μL of 2.5 mM dNTPs, 0.2 μL of 5 U / μL DNA polymerase, 0.5 μL of each 0.2 μM primer, and double-distilled water were added to 20 μL. The ITS-PCR amplification reaction conditions were: initial denaturation at 94°C for 4 minutes; 35 cycles of 94°C for 30 seconds, 56°C for 30 seconds, and 72°C for 1 minute; and extension at 72°C for 10 minutes. A 503-bp DNA fragment was amplified and sent to Sangon Biotech (Shanghai) Co., Ltd. for sequencing.
[0041] Results The nucleotide sequence of the obtained DNA fragment is shown in SEQ ID NO: 1. The sequence was compared with the NCBI database by BLAST online. The results showed that the strain sequence had the highest similarity with the ITS sequence of Phallus dongsun, with a sequence similarity of up to 96%. A phylogenetic tree was constructed. Results (see Figure 2 ) The strain Guizhou P216 of the present invention belongs to the genus Phallus dongsun, and the strain is different from any existing Phallus dongsun strain and is a new Phallus dongsun strain.
[0042] Example 3 Cultivation test of the winter iris strain Guizhou P216 of the present invention
[0043] Proceed as follows:
[0044] (1) Stock culture: Inoculate the Guizhou P216 stock culture (Guizhou P216 strain has been deposited in the General Microbiology Center of the China Microbiological Culture Management Committee, and its deposit number is: CGMCC No: 40102) onto the stock culture medium, place it in a 25°C constant temperature incubator and culture it in the dark for 20 days. The mycelium will cover the culture medium, and the Guizhou P216 stock culture will be obtained. The stock culture medium is composed of: 200g potato, 20g glucose, 20g agar, 0.3g potassium dihydrogen phosphate, 0.15g magnesium sulfate, 10mg vitamin B1, and 1000mL water. After preparation, sterilize it at 121°C for 30min and pour it into a 9cm diameter plate.
[0045] (2) Stock culture: The Guizhou P216 mother plate covered with culture medium obtained in step (1) is streaked with bacterial blocks using the well method and transferred to the stock culture medium, with one plate transferred to one bottle, and then cultured at 25°C in the dark for about 50 days until the mycelium fills the culture bottle, thus obtaining the Guizhou P216 stock. The stock culture medium comprises the following components: 89% fine sawdust, 10% wheat bran, and 1% lime; the water content is about 70%; each component is taken in proportion and mixed evenly, and then water is added in proportion; the prepared stock culture medium is placed in a 500mL glass culture bottle, sterilized under high pressure at 121°C for 1h, and placed in a clean bench to cool to room temperature after sterilization.
[0046] (3) Cultivation of cultivars: Transfer the original Guizhou P216 obtained in step (2) to a culture medium, transferring about 15 g per bag, and culturing at 25°C in the dark for about 50 days until the bag is full of fungi, thus obtaining Guizhou P216 cultivars. The culture medium comprises the following components: 80% sawdust, 18% wheat bran, 1% lime, and 1% gypsum; the water content is 65%; the components are taken in proportion and mixed evenly during preparation, and then water is added in proportion; the prepared culture medium is then placed in a polypropylene plastic bag (16 cm × 35 cm × 0.005 cm) and tied with a polypropylene rope, and sterilized at 121°C for 2 h. After sterilization, the bag is cooled to room temperature; the sawdust in the culture medium is coarse sawdust and needs to be pre-wetted in advance to prevent contamination caused by incomplete sterilization.
[0047] (4) Land preparation and bed construction: The cultivation of Guizhou P216 selects the forest cultivation mode, and the cultivation site selects a forest with a high density. The soil should be fertile and loose in texture, and the relative humidity of the air should be 70% to 85%. It is best to have a certain slope to facilitate drainage. When digging ditches, the width should be 50 cm, the depth should be 15 to 20 cm, and the length should be unlimited. The bottom of the ditch should be leveled, and 3 to 4 cm thick loose soil should be laid to retain water. The width between the two ditches should be 20 to 25 cm.
[0048] (5) Paving material: The cultivation method of Guizhou P216 is to use a layer of wood, a layer of fungus and another layer of wood. The wood is fresh broad-leaved trees, such as chestnut trees and cycads. They are cut into blocks 2 to 5 cm thick and 5 to 10 cm long. The wood is then evenly spread on the bottom of the ditch and compacted to cover the soil in the ditch.
[0049] (6) Sowing: The sowing time is April 1 to 10. Break the Guizhou P216 cultivar obtained in step (3) into small pieces with a diameter of 3 to 5 cm and sow them on the bottom wood block with a spacing of 3 to 4 cm between each strain. Then cover them with a thin layer of bamboo leaves and then a layer of wood.
[0050] (7) Covering soil: Choose clean, loose soil without obvious bacterial growth. The thickness of the covering soil should be 3 to 5 cm. In drought-prone areas, the covering soil should be slightly thicker, 5 to 8 cm. The covering soil should be in the shape of a turtle's back. Then, add a layer of pine needles about 2 cm thick to moisturize the soil.
[0051] (8) Fruiting Management: Temperature management is key during the cultivation of winter clover. Guizhou P216 is a low-temperature variety. The ideal temperature for mycelial growth is 16-24°C. The fruiting temperature for Guizhou P216 fruiting bodies is generally 5-12°C. Guizhou P216 has different humidity requirements at different growth stages. During the mycelial growth stage, the ideal humidity is 70%-85%. During the bud differentiation stage, the relative humidity is above 80%. During the fruiting body growth stage, the ideal soil humidity is 80%-85%, and the relative humidity is above 80%. Winter clover cultivation requires a semi-shaded and semi-sunny environment. Avoid direct sunlight that may damage the buds.
[0052] (9) Harvesting: The fruiting body of Guizhou P216 forms between November 1st and November 30th, and the entire growth cycle from sowing to harvesting is approximately 7 to 8 months. Winter fungus is fragile and easily broken, so be especially careful when harvesting. First, use a knife to cut off the bottom root, then gently twist and pull it up. Do not pull it directly to avoid damaging the root and affecting the secondary fruiting.
[0053] (10) After harvest, the fruiting bodies of Guizhou P216 were measured and their morphology described. The weight of each plant was weighed and the average fresh weight was calculated. The total sugar content was determined according to GB / T 15672-2009, “National Standard for Determination of Total Sugar Content in Edible Fungi.” The protein content was determined according to GB5009.5-2016, “National Food Safety Standard for Determination of Protein in Food.”
[0054] Results (see Figure 1 ) The fruiting body stipe of the winter clover strain Guizhou P216 of the present invention is white, cylindrical and hollow, with a good mushroom shape and high commercial value. Secondly, the fruiting body is large, with a stipe length of 20 to 28 cm and a diameter of 3 to 6 cm. The average weight of fresh mushrooms is 58.27 g, which is higher than the fresh mushroom weight of 25 to 40 g / piece of other existing winter clover strains. Guizhou P216 has a short cultivation cycle, and the cycle from sowing to the beginning of mushroom harvesting is 7 to 8 months, which is 2 to 4 months shorter than the cultivation cycle of 9 to 12 months of other existing winter clover strains. Guizhou P216 has a high yield. After cultivation and yield measurement, the average fresh product yield is 2.33 kg / m 2 , which is significantly higher than other existing winter iris strains by 1.0-1.5 kg / m 2 The fresh product yield is high. Testing showed that the total sugar content of Qian P216 was 28.81%, significantly higher than the 5-20% total sugar content of other existing winter iris strains. Sixth, the crude protein content of Qian P216 was 17.86%, higher than the 8-16% crude protein content of other existing winter iris strains.
[0055] The above results show that the winter iris strain Guizhou P216 of the present invention has the advantages of good mushroom shape, large fruiting body, high fresh mushroom weight, short cultivation period, high yield, and high total sugar content and crude protein content. In short, it has high nutritional value, high functional product development and utilization value, high added value, and brings high economic benefits to mushroom farmers.
Claims
1. A kind of winter clover ( Phallus dongsun ) strain Guizhou P216 was deposited in the General Microbiology Center of China Culture Collection Administration of Microorganisms, and its deposit number is CGMCC No.40102.
2. Use of the strain Guizhou P216 according to claim 1 in food.
3. A food, characterized in that The food contains the strain Guizhou P216 according to claim 1.
4. Use of the strain Guizhou P216 according to claim 1 in beverages.
5. A beverage, characterized in that The beverage contains the strain Guizhou P216 according to claim 1.