Schizophyllumcommuneh spaceflight-carried schizophyllumcommuneh strain

Abstract

The invention relates to a spaceflight-carried schizophragma integrifolium strain Kungsu No.3 and application thereof, and belongs to the technical field of strain breeding. The spaceflight-carried schizophyllum commune strain No.3 is preserved in the China General Microbiological Culture Collection Center (CGMCC) on August 9, 2024, the preservation number is CGMCC No.41428, the classification name of the spaceflight-carried schizophyllum commune strain is schizophyllum commune, and the preservation address is No.3, Yard 1, Beichen West Road, Chaoyang District, Beijing. Compared with cultivation of strains which are not subjected to spaceflight carrying, the Kunzhou No.3 strain is fast in hypha germination, fast in growth, crisp in sporocarp, free of or few in spores, light pink to pink, high in conversion rate, short in period and suitable for industrial cultivation.

Description

Technical Field

[0001] The present application relates to the technical field of bacterial strain selection and breeding, and in particular to a space-borne white ginseng bacterial strain Kunzhi No. 3 and its application. Background Art

[0002] White ginseng is the trade name of Schizophyllum, which belongs to the order Agaricales, family Schizophyllum, genus Schizophyllum. Wild Schizophyllum is tender and delicious when young, with a special strong aroma, and is a famous edible fungus in Yunnan. Domestic and foreign medical research shows that the fruiting bodies of Schizophyllum contain organic acids and Schizophyllum polysaccharides. The fruiting bodies of Schizophyllum are small, have many spores, and are tough (commonly known as Badanchai), and the biological conversion rate of artificial cultivation is low.

[0003] The efforts to breed sporeless or oligospore white ginseng and improve the taste and yield of white ginseng have always been pursued by those skilled in the art, and have great application value. Summary of the invention

[0004] The present application provides a white ginseng strain Kunzhi No. 3, which has the characteristics of fast mycelium germination, fast growth, brittle fruiting body, no or few spores, light pink to pink color, high conversion rate and short cycle.

[0005] The white ginseng strain Kunzhi No. 3 in this application was deposited in the General Microbiological Center of China Microbiological Culture Collection Administration on August 9, 2024, with the deposit number CGMCC No.41428, and the classification name was Schizophyllum ( Schizophyllum commune ), the storage address is No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing.

[0006] Specifically, the new white ginseng strain Kunzhi No. 3 in the present application is obtained by the following breeding method: First, a series of strains were obtained by tissue separation and purification from wild Schizophyllum fruiting bodies collected in Lulang Town, Nyingchi City, Tibet Autonomous Region in August 2019. Then, the strain to be carried in space (No. S12-1) was selected through primary screening, re-screening, regional trials and production trials.

[0007] On June 17, 2021, the white ginseng strain numbered S12-1 was carried into space by the Shenzhou XII manned spacecraft, and returned to the ground on September 17, 2021 with the Shenzhou XII manned spacecraft; the S12-1 strain was activated, the activated mycelium was ground, and the single root fragments were separated and cultured to obtain S12-1~200; further screening was carried out according to the conventional breeding methods for edible fungi to obtain an excellent strain with fast mycelium germination, brittle fruiting body, no or few spores, light pink to pink color, high yield and short cycle, numbered Kunzhi No. 3.

[0008] The present application also provides an application of the white ginseng strain Kunzhi No. 3 in the cultivation and production of white ginseng. Applying the strain Kunzhi No. 3 provided in the present application to the factory production of white ginseng can reduce the impact of spores on the environment and workers, improve the conversion rate of white ginseng, and has broad application prospects.

[0009] The beneficial effects of this application are: Compared with the strain S12-1 that was not cultivated on space flight, the mycelium of the Kunzhi No. 3 strain germinated quickly, the fruiting body was brittle, there were no or few spores, the color was light pink to pink, the yield was high and the cycle was short, which has great application value. BRIEF DESCRIPTION OF THE DRAWINGS

[0010] The above and other objects, features and advantages of the present application will become more apparent through a more detailed description of exemplary embodiments of the present application in conjunction with the accompanying drawings, wherein the same reference numerals generally represent the same components in the exemplary embodiments of the present application.

[0011] Figure 1 This is a schematic diagram of the morphology of the strain Kunzhi No. 3 obtained in Example 1 of the present invention; Figure 2 This is a schematic diagram of the strain Kunzhi No. 3 in Example 2 of the present invention being cultivated and about to produce mushrooms. DETAILED DESCRIPTION

[0012] The embodiments of the present application will be described in more detail below with reference to examples. Although the embodiments of the present application are shown in the examples, it should be understood that the present application can be implemented in various forms and should not be limited by the embodiments set forth herein. On the contrary, these embodiments are provided to make the present application more thorough and complete, and the scope of the present application can be fully communicated to those skilled in the art.

[0013] Example 1 This example illustrates the breeding process of white ginseng strain Kunzhi No. 3, and the specific steps are as follows: (1) A series of strains were obtained by tissue separation and purification from wild Schizophyllum sporocarp fruiting bodies collected in Lulang Town, Nyingchi City, Tibet Autonomous Region in August 2019. Then, a strain (numbered S12-1) was selected for space travel after primary screening, secondary screening, regional testing and production testing. The strain was amplified and identified using universal fungal ITS primers (ITS4 / ITS5), and the identification result was Schizophyllum sporocarp.

[0014] (3) The white ginseng strain numbered S12-1 was carried into space on the Shenzhou XII manned spacecraft on June 17, 2021. It returned to the ground in the Shenzhou XII manned spacecraft on September 17, 2021. After notarization, it was brought back to the Culture Collection Center of the Kunming Institute of Botany, Chinese Academy of Sciences for relevant research.

[0015] (4) The space-borne strain S12-1 was activated, the activated mycelium was ground, and the single root fragments were separated and cultured to obtain S12-1~200.

[0016] (5) With reference to GB / T 21125-2007 Technical Specifications for Breeding of Edible Fungi, further screening was carried out according to conventional breeding methods for edible fungi, and an excellent strain with fast mycelial growth, crisp fruiting body, light pink to pink color, no or few spores, high conversion rate and short cycle was obtained, which was numbered Kunzhi No. 3. The strain was preserved in the Kunming Institute of Botany, Chinese Academy of Sciences (KUNCC) using the 10% glycerol + distilled water preservation method.

[0017] (6) The white ginseng strain Kunzhi 3 was deposited in the General Microbiology Center of China Microorganism Culture Collection Administration on August 9, 2024, with the deposit number: CGMCC No.41428, and the classification name is Schizophyllum ( Schizophyllum commune ), the storage address is No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing.

[0018] The characteristics of the white ginseng strain Kunzhi No. 3 in this embodiment are as follows: 1. Morphological characteristics The fruiting body has a cap with a diameter of 2-7 cm, light pink to pink, with velvet or coarse hairs, fan-shaped or kidney-shaped, with many lobes, rolled inward; the flesh is crisp, white with a slight pink tint; the gills are narrow, radiating from the base, light pink to pink, and longitudinally cracked along the edges; the whole plant is chrysanthemum-shaped. Figure 1 shown.

[0019] 2. Biological characteristics 1) Mycelium culture: After inoculation, the cultivation bag is sealed with conventional edible mushroom tape and cultured in a dark environment at an air temperature of 25°C and an air humidity of 65% for 7 to 10 days to prepare for fruiting.

[0020] 2) Management of mushroom production: After 7-10 days of mycelium cultivation, remove the sealing glue, maintain the temperature of the mushroom house at 16-21°C, the relative humidity of the air at 90-92%, the carbon dioxide concentration at 450-550ppm, the light intensity at 250lx, the light at 20min / h, and harvest in time when the fruiting body diameter reaches 5-15cm.

[0021] 3. Genetic characteristics The genomic DNA of the white ginseng strain Kunzhi 3 was first amplified and sequenced using the universal primers ITS4 / ITS5 for fungi for the internal transcribed spacer region (ITS) of the ribosome. The DNA sequence bases obtained by sequencing are shown in SEQ ID NO.3, specifically: CATCTTGTTCTGATCCTGTGCACCTTATGTAGTCCCAAAGCCTTCACGGGCGGCGGTTGACTACGTCTACCTCACACCTTAAAGTATGTTAACGAATGTAATCATGGTCTTGACAGACCCTAAAAAGTTAATACAAC TTTCGACAACGGATCTCTTGGCTCTCGCATCGATGAAGAACGCAGCGAAATGCGATAAGTAATGTGAATTGCAGAATTCAGTGAATCATCGAATCTTTGAACGCACCTTGCGCCCTTTGGTATTCCGAGGGGCATGCC TGTTTGAGTGTCATTAAATACCATCAACCCTCTTTTGACTTCGGTCTCGAGAGTGGCTTGGAAGTGGAGGTCTGCTGGAGCCTAACGGAGCCAGCTCCTCTTAAATGTATTAGCGGATTTCCCTTGCGGGATCGCGT CTCCGATGTGATAATTTCTACGTCGTTGACCATCTCGGGGCTGACCTAGTCAGTTTCAATAGGAGTCTGCTTCTAACCGTCTCTTGACTGAGACTAGCGACTTGTGCGCTAACTTTTGACTTGACCTCAAATCAGGTA The fragment was sequenced by GenBank, and the result showed 100% homology with the white ginseng in the database, gene accession number (OR198218). The white ginseng strain Kunzhi 3 (preservation number CGMCC No. 41428) was identified as white ginseng.

[0022] Subsequently, the genome of the white ginseng strain Kunzhi 3 was sequenced using a universal fungal genome sequencing method, and shallow genome sequencing (Shallow Whole Genome Sequencing) and primer design were performed with reference to existing technologies to obtain the following specific primers: Primer_KZ3_1L: 5'-ATTCGGTCGAGATGGTGAAC-3' Primer_KZ3_1R: 5'- TGTTGTCTGCTATCCCAATGC-3' Primer_KZ3_2L: 5'-TTTCGAGCGTTTCCTTCCAT-3' Primer_KZ3_2R: 5'- CTTGTCAATCTGCGGGGAC-3' Specific gene detection was performed on the strain Kunzhi No. 3 selected after the spacecraft was carried on board the Shenzhou XII-IV manned spacecraft, the strain not carried on board, and the common white ginseng on the market. During amplification, Primer_KZ3_1L / Primer_KZ3_1R or Primer_KZ3_2L / Primer_KZ3_2R primers were added to the corresponding amplification system respectively, and the reaction was detected by 2% agarose gel electrophoresis after the reaction was completed.

[0023] The amplification system included: PCR Buffer 1×, MgCl2 1.50 mM, dNTPs Mix 0.25 mM, primers 1.00 mM each, rTaq Polymerase 1.5 U, and template DNA 50 ng; the reaction conditions were: pre-denaturation at 95°C for 3 min, followed by 95°C for 60 s, 58°C for 40 s, and 72°C for 30 s, for 35 cycles; and finally extension at 72°C for 10 min.

[0024] The sequencing of the PCR amplification reaction products was sent to Beijing Biotech Biotechnology Co., Ltd., Shanghai Shenggong Bioengineering Technology Service Co., Ltd. and Yingjun Biotechnology Co., Ltd. The sequencer used for sequencing was ABI3730 DNA sequencer, and the sequencing primers were the same as the PCR amplification primers.

[0025] Specific DNA fragments of about 128 bp and 238 bp were obtained by PCR amplification, and the DNA sequence bases obtained by sequencing are shown in SEQ ID NO.1 and SEQ ID NO.2, respectively. Specifically, SEQ ID NO.1 is: TTAGTCCCGTAGAGGACGATTCGGTCGAGATGGTGAACTCGAAGCCCGACGAAGCGACGTCCTCGATGGTGATGCTAGCGCTGAGCGTAATGGACGCTGGAAGGTACGTGACCACCGCGTCGCGTCGCATGGGATAGCAGACAACATACCACTGTTGGTCACGGTGAGGGTGGCGGAGTTCGTGTTGGTCCACGACTCCG Specifically, SEQ ID NO.2 is: GAACCAAACTCTCTGCCGTTTCGAGCGTTTCCTTCCATAACTTTTGCTCCAGCGCACGGCCGGCGGGTGTCTCCGAGAATGGGAAGGGCCTAATAGAGGTCAGATAAGAGTTTGCGAGGGAATTCAGACGAAAGAC GCACGGGAAAACCTCCGTGTCTGCAATGAGTGCCCCGTGGCTATCCGACCGTGTGGTGGTAGCAGCGTATACGACACAGCGTGCGCCTTCCTCGGGAGTACGTCCCCGCAGATTGACAAGCGGCCTGATGAGCGAG Only the Kunzhi No. 3 strain can obtain the above DNA sequence bases. The uninfected strains or the common white ginseng in the market cannot amplify the above DNA sequence bases.

[0026] Only the Kunzhi No. 3 strain can amplify the unique DNA fragment using the specific primers.

[0027] Example 2 Cultivation Comparative Test In this example, the white ginseng strain Kunzhi No. 3 was cultivated and compared with the strain S12-1 that was not carried on space flight, and the mycelial growth rate, spore number, growth cycle and yield per unit area of ​​the original strain and the cultivated strain of the test strain were examined.

[0028] The mother culture formula used in this embodiment is PDA culture medium commonly used for edible fungi.

[0029] The stock formula used in this embodiment is a common stock culture medium for wood-decaying fungi: 68% wheat grains, 30% sawdust, 1% light calcium carbonate, 1% gypsum, 58% water content, pH 6 after sterilization; The cultivation formula used in this embodiment is a common culture medium for wood-decaying fungi: 50% sawdust, 20% cottonseed hulls, 10% wheat bran, 18% corn flour, 1% light calcium carbonate, 1% gypsum, with a water content of 60-63% and a pH of 6 after sterilization.

[0030] Furthermore, the cultivation method of the Kunzhi No. 3 strain in this embodiment comprises the following steps: 1) Preparation of mother species, original species and cultivated species: According to the general preparation method of wood-decaying fungi, the above formula is prepared into corresponding mother species, original species and cultivated species for later use.

[0031] 2) Mycelium culture: After inoculation of the mother culture, original culture and cultivated culture, culture them in a dark place at an air temperature of 25°C and an air humidity of 65% for 10 to 15 days.

[0032] 3) Mushroom management: After 7-10 days of cultivation of mycelium, remove the sealing glue, maintain the temperature of the mushroom house at 16-21°C, relative humidity at 90-92%, carbon dioxide concentration at 450-550ppm, light intensity at 250lx, light at 20min / h, and when the fruiting body diameter reaches 5-15cm (such as Figure 2 As shown), harvest in time.

[0033] It should be noted that in Example 2, when comparing the cultivation of Kunzhi No. 3 and S12-1, the only difference was the strain itself, and the other cultivation conditions were the same.

[0034] The cultivation results are shown in the following table: It can be seen from the above table that compared with the strain S12-1 that was not cultivated by space transport, the mycelium of the Kunzhi No. 3 strain grew faster in the cultivation bag, the fruiting body was brittle, there were no or few spores, and the color was light pink to pink, and the conversion rate and yield were significantly improved.

[0035] The embodiments of the present application have been described above, and the above description is exemplary, not exhaustive, and is not limited to the disclosed embodiments. Many modifications and changes will be apparent to those of ordinary skill in the art without departing from the scope and spirit of the described embodiments. The selection of terms used herein is intended to best explain the principles of the embodiments, practical applications, or improvements to the technology in the market, or to enable other persons of ordinary skill in the art to understand the embodiments disclosed herein.

Claims

1. A space-borne white ginseng strain Kunzhi No. 3, characterized in that: It was deposited in the General Microbiology Center of China Microbiological Culture Collection Administration on August 9, 2024, with the deposit number CGMCC No.41428, and the classification name was Schizophyllum ( Schizophyllum commune ), the storage address is No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing.

2. An application of the strain Kunzhi No. 3 according to claim 1 in white ginseng production.

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