CN103503779A - New pure white hypsizigus marmoreus strain - Google Patents
New pure white hypsizigus marmoreus strain Download PDFInfo
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Abstract
The invention provides a new pure white hypsizigus marmoreus strain Finc-W-227 of which the collection number is CCTCC NO:M2012377. The new pure white hypsizigus marmoreus strain Finc-W-227 has the advantages of low probability of occurrence of cap mushroom tumors, and high single yield, and is suitable for large-area popularization and planting.
Description
[technical field]
The present invention relates to edible mushroom technical field, more precisely, relate to the new bacterial strain of the true Ji mushroom of a kind of pure white.
[background technology]
True Ji mushroom (Latin title Hypsizygus marmoreus), is subordinate to Basidiomycotina, Hymenomycetes, Agaricales ,Bai Mo Ke,Li gill fungus family, beautiful gill fungus genus, because it has unique crab delicate flavour, therefore claim that again it is hypsizygus marmoreus.Initial artificial cultivation started from Japan before and after 1978, and its delicious flavour, sense of food are tender and crisp, nutritious, and its taste is brighter than flat mushroom, and its meat is thicker than sliding mushroom, and its matter is more tough than mushroom, and mouthfeel is excellent, and its dry product also thick flavor overflows.True Ji mushroom contains enriches vitamin and 17 seed amino acids, β-1 of extracting in fruit body, and 3-D glucan has very high antitumor activity, is a kind of health food low in calories, low-fat.
True Ji mushroom is from cultivating beginning, and what substantially walk is exactly the route that batch production is produced.Because factory culture has the anniversary, produce continuously, large-scale production, mostly realize semi-mechanization, mechanization so that intelligent, growing environment is controlled better, yield and quality has had good assurance.As everyone knows, bacterial classification plays vital effect to the factory culture of edible mushroom.Current true Ji mushroom has dark brown and two main cultivation strains of pure white, and pure white strain claims again " white jade mushroom ", " Yulong mushroom ", is received by the market, and main breed only has the Big Dipper white jade mushroom No. 1 (hereinafter to be referred as " H-W ") of Japanese Big Dipper company.H-W is the true Ji mushroom of the pure white strain that seed selection in 2002 is released, and its entire body is snow-white, without bitter taste, increased the aesthetic feeling of dish, but it still exists following defect: knurl lid mushroom easily appears in (1), and yields is low; (2) per unit area yield is lower, affects economic benefit.
[summary of the invention]
The technical problem that the present invention solves is to overcome the defect that prior art exists, and provides a kind of per unit area yield high, and the low new bacterial strain Finc-W-227 of the true Ji mushroom of pure white of knurl lid mushroom occurrence rate.
The present invention is achieved by the following technical solutions, and the new bacterial strain Finc-W-227 of the true Ji mushroom of pure white of the present invention is by after parent TNN-11 and H-W hybridization, then obtains through systematic breeding.This bacterial strain has been preserved in Chinese Typical Representative culture collection center, address on September 24th, 2012: Wuhan, China city Wuhan University, and its preserving number is CCTCC NO:M2012377, its biology Classification And Nomenclature is Hypsizygus marmoreus Finc-W-227.
The new bacterial strain Finc-W-227 of the true Ji mushroom of pure white of the present invention is characterised in that:
1, morphological feature
The mycelia of Finc-W-227: pure white sparse, aerial hyphae is undeveloped; Cap: section Cheng Yuan mountain type, entire body is snow-white, and meat is thick, meat is crisp, and speckle is clear, central distribution, without be full of cracks, not rounding; Lamella: snow-white, wavy bending, sparse; Without spore print; Stem: thick, snow-white, nothing hair, slightly eccentric raw with cap, meat is crisp.
2, biological property
Nutrient component source is: corncob, weed tree sawdust, rice bran, wheat bran, corn flour, cotton seed hulls etc., wherein, water content 64～66%.20～25 ℃ of Mycelium culture temperature, humidity 75%, CO2 concentration 2500～4000ppm, cultivation cycle 70～90d.Suitable fruiting temperature is 13～15 ℃, humidity 93～100%, and CO2 concentration 1000～2000ppm, front 1～7d intensity of illumination 50～100Lux, rear 8～20d intensity of illumination 200～300Lux, every day, clearance-type was opened layer bracket lamp stimulation fruiting, cultivation cycle 24～26d.
Compared with prior art, the new bacterial strain Finc-W-227 of the true Ji mushroom of pure white of the present invention has reduced knurl lid mushroom probability of occurrence effectively, has improved single rate simultaneously, suits large area to popularize.
[accompanying drawing explanation]
Fig. 1 is that Finc-W-227 and H-W per unit area yield, cultivation cycle are with cell age change curve.
Fig. 2 is that Finc-W-227 and H-W mycelial growth rate vary with temperature curve map.
Fig. 3 is primer I TS1 and the pcr amplification electrophoresis pattern of ITS4 to 4 bacterial strains, and wherein 1 is bacterium H-W, and 2 is Finc-W-227, and 3 is G-W, and 4 is GC-W, and M is 10kbp DNA ladder.
Fig. 4 is Finc-W-227 phylogenetic tree.
Fig. 5 is the pcr amplification electrophoresis pattern of primer S99 to 4 bacterial strains, numbers same Fig. 3.
[embodiment]
Described " G-W " bacterial strain in following examples is a kind of true Ji's mushroom bacterial strain of buying from the market, and its former base is grey, and along with the g and D of fruit body bleaches gradually, picking time, stem was canescence, and cap is pure white; Described " GC-W " bacterial strain is the new white jade mushroom bacterial strain being bred as of Japanese Ge Cheng.
The breeding process of the new bacterial strain Finc-W-227 of the true Ji mushroom of pure white of the present invention comprises the steps:
1) Selection parent bacterial strain: choosing TNN-11 and H-W is parent, on these two parents' fruit body Jun Neng market, purchase obtains ， Academy of Agricultural Sciences, Shanghai City DSMZ also preservation.
2) preparation medium
PDA medium: every 1L medium potato 200g, glucose 20g, agar 20g.
Pedigree seed culture medium: wood chip 65%(percentage by weight, lower with), rice bran 18%, wheat bran 7%, corn flour 10%, water content is 60%～62%.
Cultivated species medium: corncob 21%, weed tree sawdust 15%, rice bran 20%, wheat bran 20%, corn flour 5%, cotton seed hulls 19%, wherein water content 64%～66%.
3) conventional method obtains 800 of heterozygotes.
4) cultivation screening: the heterozygote obtaining is carried out to experiment in cultivation and screening according to conventional method.
5) obtain aimed strain: by screening system, from 800 heterozygotes, obtain a pure white bacterial strain, its production capacity is high, has improved productivity effect, has reached breeding goal.This bacterial strain is named as " Finc-W-227 ".
The new bacterial strain Finc-W-227 of the true Ji mushroom of pure white of the present invention is preserved in Chinese Typical Representative culture collection center on September 24th, 2012, and deposit number is CCTCC NO:M2012377, and bacterial strain is survived after testing, and classification position is that beautiful gill fungus belongs to true Ji mushroom.
The present embodiment by plantation in the new bacterial strain Finc-W-227 of the true Ji mushroom of pure white of the present invention and the true Ji mushroom of existing pure white the most widely H-W to carry out cultivation cycle (be cell age, lower with), the contrast of cultivation cycle and per unit area yield, advantage and the feature of the new bacterial strain Finc-W-227 of the true Ji mushroom of pure white of the present invention are described.
The cultivated species medium that the present embodiment adopts is: corncob 35%, weed tree sawdust 14%, rice bran 14%, wheat bran 19%, and corn flour 5%, cotton seed hulls 13%, wherein water content 64%～66%.
The cell age test of table 1 Finc-W-227 and H-W
Take H-W as contrast, is sky from 70d(, lower same) to 105d, 8 cell ages are set, every increase 5d is a cell age, 80 bottles of every cell ages.Cultivate and cultivate the true Ji mushroom cultivation and the cultivation parameter that adopt routine, the volume of culture bottle is 850cc, bottleneck diameter 58mm.
This result of the test sees also Fig. 1 and table 1, and the highest per unit area yield of H-W is that 161g/bottle(is gram/bottle, lower same), best cell age is 95d, and the corresponding cultivation cycle is 23d, and the highest per unit area yield of Finc-W-227 reaches 228g/bottle, best cell age is 90d, and the cultivation cycle is 25d.Known by contrasting, Finc-W-227 advantage aspect single rate is very obvious, in the scope of cell age at 70d～90d, the cultivation cycle of Finc-W-227 is in 25d left and right, the cultivation period ratio H-W of 75d and two cell age Finc-W-227 of 80d shortens, and the cultivation period ratio H-W of the Finc-W-227 of 85d and two cell ages of 90d is slightly long.Described in comprehensive, in 70d～105d cell age, the single rate of Finc-W-227 improves 30.6%～45.2% than H-W.
The fresh-keeping test of embodiment 3
Take H-W as contrast, after Finc-W-227 and H-W gather, by every bag 155～165g, pack, each 20 bag, in freezer (3 ℃～8 ℃) preservation 11d, then takes out and is placed under 23 ℃ of room temperature conditions, records Fruitbody situation of change.Result of the test refers to table 2, and after freezer (3 ℃～8 ℃) preservation 11d, it is still intact that Finc-W-227 continues preservation 11d at 23 ℃ of environment; The 10d that H-W continues preservation at 23 ℃ of environment starts, by snow-white flavescence, to have lost feeling of freshness, and Finc-W-227 13d under identical environment partly starts flavescence; H-W is after freezer preservation 11d, and 23 ℃ of 18d that continue preservation all lose commercial value, and Finc-W-227 continues just all to lose commercial value after preservation 20d at 23 ℃.The freshness date that Finc-W-227 is more than described slightly has superiority than H-W.
The fresh-keeping test of table 2 Finc-W-227 and H-W
The inoculation piece that is 5.0mm Finc-W-227 and H-W diameter is inoculated on PDA medium, cultivates 11d respectively at 20 ℃, 25 ℃, 30 ℃ and 35 ℃, and result of the test is in Table 7 and Fig. 2.Result of the test shows: the mycelia of Finc-W-227 is very fast, visible 20 ℃～25 ℃ growth rate, and its suitable cultivation temperature is 20～25 ℃.
The mycelial growth temperature gradient test of table 3 Finc-W-227 and H-W
Embodiment 5 morphological features and biological property
Morphological feature: Finc-W-227 mycelia is pure white sparse, and aerial hyphae is undeveloped; Cap section circle mountain type, entire body is snow-white, and meat is thick, meat is crisp, and speckle is clear, central distribution, without be full of cracks, not rounding; Snow-white, the wavy bending of lamella, sparse; Without spore print; Stem is thick, snow-white, nothing hair, and slightly eccentric raw with cap, meat is crisp.The difference of the concrete form feature of Finc-W-227 and H-W refers to table 4.
Table 4 Finc-W-227 and the contrast of H-W morphological feature
Biological property: nutrient component source is: corncob, weed tree sawdust, rice bran, wheat bran, corn flour and cotton seed hulls etc., wherein, water content 64%～66%.20～25 ℃ of Mycelium culture temperature, humidity 75%, CO
2concentration 2500ppm～4000ppm, cultivation cycle 70d～90d.Suitable fruiting temperature is 13～15 ℃, humidity 93%～100%, CO
2concentration 1000ppm～2000ppm, front 1d～7d intensity of illumination 50Lux～100Lux, rear 8d～20d intensity of illumination 200Lux～300Lux, every day, clearance-type was opened layer bracket lamp stimulation fruiting, cultivation cycle 24d～26d.
Embodiment 6 antagonistic effects
It is test material that the present embodiment be take the new bacterial strain Finc-W-227 of the true Ji mushroom of pure white of the present invention and H-W, G-W, GC-W, and used medium is PDA medium.
Finc-W-227 and H-W, G-W, GC-W be face-off inoculation and cultivation respectively on plating medium, and vaccination is at a distance of 2cm, 24 ℃ of constant temperature culture 26d.
Antagonistic effect result is with reference to table 5, in face-off, cultivate the front and back of bacterium colony intersection, Finc-W-227 and H-W, Finc-W-227 and G-W and Finc-W-227 and GC-W antagonism line are obvious, illustrate that Finc-W-227 is the true Ji mushroom new bacterial strain different with GC-W from H-W, G-W.
Antagonistic effect between the different true Ji mushrooms of table 5
Embodiment 7 ITS order-checkings are identified
In the present embodiment, ITS order-checking student on commission's work biotechnology (Shanghai) limited company (hereinafter to be referred as " raw work ") carries out, and comprises following process:
Genomic extraction: extract according to raw work SK1375 fungal genomic DNA extraction agent box specification.Resulting DNA solution is placed in to-20 ℃, refrigerates standby.
Pcr amplification and Establishing (25 μ L): genomic templates 1 μ L, fungi rDNA universal primer ITS1 5'-TCCGTAGGTGAACCTGCGG-3'(10 μ M) 0.5 μ L, ITS4 5'-TCCTCCGCTTATTGATATGC-3'(10 μ M) 0.5 μ L, dNTP mix (10mM each) 0.5 μ L, 10 * Taq reaction buffer, 2.5 μ L, Taq (5u/ μ L) 0.2 μ L, adds water to 25 μ L.PCR program: 94 ℃ of 5mim of denaturation; 94 ℃ of 30s, 55 ℃ of 35s, 72 ℃ of 1min, 35 circulations, 72 ℃ are extended 8min.The PCR product that amplification obtains is by agarose gel electrophoresis, and its electrophoresis result is shown in Fig. 3.
DNA Ago-Gel reclaims glue purification and order-checking adopts conventional method.
The ITS sequence sequencing result of this test strain Finc-W-227 is shown in sequence table, the DNA fragmentation that its sequence is 665bp.From GenBank, check in, the ITS sequence similarity that Finc-W-227 bacterial strain ITS sequence and beautiful gill fungus belong to true Ji mushroom is the highest.And from GenBank, checking in the ITS sequence that belongs to the close kind of true Ji mushroom with beautiful gill fungus, application DNAStar MegAlign software building goes out the phylogenetic tree of strains tested, refers to Fig. 4.
Embodiment 8 RAPD analyze
It is analytic target that the present embodiment be take the new bacterial strain Finc-W-227 of the true Ji mushroom of pure white of the present invention and H-W, G-W, GC-W, utilizes RAPD analytical technology, further identifies the hereditary capacity of the new bacterial strain of the true Ji mushroom of pure white of the present invention.In the present embodiment, DNA extraction, pcr amplification and agarose gel electrophoresis are identical with embodiment 7, do not repeat them here.
Refer to table 6, during RAPD analyzes, filter out 1 primer and can distinguish clearly Finc-W-227 and H-W, G-W, the true Ji's mushroom strains of GC-W from 12 random primers, this random primer is S99.
Table 6 random primer
The result that RAPD analyzes is as follows:
The pcr amplification agarose gel electrophoretogram that primer S99 is corresponding is Fig. 5, the position at a frame place, and Finc-W-227 is without DNA band, and H-W, G-W and GC-W have DNA band, illustrate that Finc-W-227 and H-W, G-W and GC-W are different bacterial strains.More than illustrate that primer S99 can distinguish Finc-W-227 and H-W, G-W and GC-W bacterial strain, Finc-W-227 is the new bacterial strain of true Ji mushroom.
More than describing is only embodiments of the invention, forgives and can understand, and is not departing under the prerequisite of the present invention's design, within all should being included in technical conceive of the present invention to simple modification of the present invention and replacement.
Claims (1)
1. the new bacterial strain Finc-W-227 of the true Ji mushroom of pure white, its deposit number is CCTCC NO:M 2012377.
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| CN105165635A (en) * | 2015-10-22 | 2015-12-23 | 青岛丰科生物科技有限公司 | Hypsizygus marmoreus strain and application thereof |
| CN106434381A (en) * | 2016-10-31 | 2017-02-22 | 上海丰科生物科技股份有限公司 | Pure white hypsizigus marmoreus strain, and molecular marker, specific primer pair and application thereof |
| CN113637591A (en) * | 2020-04-27 | 2021-11-12 | 西充星河生物科技有限公司 | Hypsizigus marmoreus strain |
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| CN105165635A (en) * | 2015-10-22 | 2015-12-23 | 青岛丰科生物科技有限公司 | Hypsizygus marmoreus strain and application thereof |
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| CN106434381A (en) * | 2016-10-31 | 2017-02-22 | 上海丰科生物科技股份有限公司 | Pure white hypsizigus marmoreus strain, and molecular marker, specific primer pair and application thereof |
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| CN113637591A (en) * | 2020-04-27 | 2021-11-12 | 西充星河生物科技有限公司 | Hypsizigus marmoreus strain |
| CN113637591B (en) * | 2020-04-27 | 2023-10-03 | 西充星河生物科技有限公司 | Hypsizygus marmoreus strain |
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