CN102907257A - Cultivation method of hazel oudemansiella mucida and special culture medium thereof - Google Patents
Cultivation method of hazel oudemansiella mucida and special culture medium thereof Download PDFInfo
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Abstract
The invention discloses a cultivation method of a hazel oudemansiella mucida and a special culture medium thereof. The cultivation method of the hazel oudemansiella mucida comprises the step of cultivating the hazel oudemansiella mucida with a sporocarp culture medium, wherein the sporocarp culture medium is a culture medium formed by mixing a cotton seed hull, apple branch sawdust, wheat bran, a hydrated lime and water and the water content of the culture medium is 65%-70% (mass percent); and the mass ratio of the cotton seed hull to the apple branch sawdust to the wheat bran to the hydrated lime is 50:30:18:2. The biology efficiency of the cultivation method of the hazel oudemansiella mucida is up to 109%-123%; the time of inserting a strain in the sporocarp culture medium till a first batch of the hazel oudemansiella mucida is harvested is 31-42 days; and the whole cultivating production cycle of harvesting four batches of the hazel oudemansiella mucida is totally 55-79 days.
Description
Technical field
The present invention relates to cultivation method and the special culture media thereof of a kind of edible mushroom, particularly the cultivation method of light brown Aode mushroom and special culture media thereof.
Background technology
Edible mushroom light brown Aode mushroom (Oudemansiella canarii) claims again Oudemansiella canarii, is under the jurisdiction of white mushroom section (Tricholomataceae) Genus Oudeman Siella Spec (Oudemansiella) of Agaricales (Agaricales).Its fruit body is general medium large.Bacteria cover diameter 3-10cm, surperficial brown is to sepia, and is sticking when moistening.Bacterial context white.Lamella white is to dirty white, and is rarer, not isometric, straight giving birth to prolonging life, and the coarse brown that is of pleat edge is to dark-coloured, and stem is often crooked, dirty white, there are dark brown cilium and vertical stripe in the surface, and is inner soft to becoming hollow.Spore oval, wide ovum are justified to subsphaeroidal, 12-23 μ m * 10.5-18 μ m.Pleat edge utricule reaches 80-150 μ m, the wide 12-40 μ m that reaches.Summer, autumn is scattered or singly be born in the deciduous forest on the rotten wood.Mainly be distributed in the ground such as Yunnan, Hainan, Heilungkiang, Jilin in China.
At present, the source of light brown Aode mushroom (Oudemansiella canarii) mainly is from field acquisition.Domestic part bacterial strain to Genus Oudeman Siella Spec has been realized artificial domesticating cultivation, such as long root oudemansiella radicata (O.radicata), Oudemansiella furfura (O.furfuracea), brown ruffle oudemansiella radicata (O.brunneomarginata), but there are no the report of artificial cultivation light brown Aode mushroom (Oudemansiella canarii); External Marcelo Jos é Silveira Ruegger etc. discloses the method for a kind of cultivation light brown Aode mushroom (Oudemansiella canarii), the method wheat bran and bagasse (mass ratio 1:4) or wheat bran and eucalyptus wood chip (mass ratio 1:4) cultivation light brown Aode mushroom, its the highest biological efficiency is for adopting the culture medium for cultivating of wheat bran and bagasse, concrete numerical value is 55.66%(Marcelo Jos é SilveiraRuegger, et al.CULTIVATION OF THE EDIBLE MUSHROOM OUDEMANSIELLACANARII (JUNGH.)
IN LIGNOCELLULOSIC SUBSTRATES.Brazilian Journalof Microbiology (2001) 32:211-214).Carry out biological property and the artificial cultivation research of light brown Aode mushroom, the cultivation method that filters out the light brown Aode mushroom (O.canarii) of higher biological efficient will effectively reduce the production cost of light brown Aode mushroom, increase mushroom farming income, and can be this artificial domesticating cultivation that belongs to other kinds and provide fundamental basis and practical experience, have important application value.
Summary of the invention
An object of the present invention is to provide the cultivation method of the higher light brown Aode mushroom of a kind of biological efficiency.
The cultivation method of light brown Aode mushroom provided by the present invention, comprise the step with fruit body medium culture light brown Aode mushroom, described fruit body medium is that cotton seed hulls, apple branch wood chip, wheat bran, white lime and water are mixed and made into water content is 65%-70%(quality percentage composition) medium, wherein, the proportioning of cotton seed hulls, apple branch wood chip, wheat bran, white lime is 1)-5) in any:
1）50：30：18：2；
2）42：38：18：2；
3）（42-50）：（30-38）：18：2；
4）（50-65）：（15-30）：18：2；
5）42-65：15-38：18：2。
Wherein, the C/N ratio of described fruit body medium can be 50-60,56-60,50-56,56,60 or 50.
Wherein, described cultivation method specifically comprises the steps:
1) send out bacterium: light brown Aode mushroom (Oudemansiella canarii) second class inoculum is accessed described fruit body medium, is 24 ℃-26 ℃ in environmental temperature, and relative air humidity is to be cultured to mycelia under the condition of 50%-60% to cover with medium;
2) low temperature stimulation: the culture of step 1) is transferred under-5 ℃ of-5 ℃ of conditions processed 3-5 days, finish low temperature stimulation;
3) fruiting; It is 23 ℃-26 ℃ that the culture of step 1) is transferred to environmental temperature, relative air humidity 80%-90%, intensity of illumination 300-600Lux(such as 300Lux or 600Lux), gas concentration lwevel is 1500-2000ppm(ppm: 1,000,000 air molecules have a carbon dioxide molecule) be cultured to fruit body primordium under the condition of (such as 1500ppm or 2000ppm) and form.
In the above-mentioned cultivation method, described second class inoculum is that female kind of light brown Aode mushroom (Oudemansiella canarii) accessed the culture that the second class inoculum medium obtains; Described second class inoculum medium can be apple branch wood chip, wheat bran, sucrose, white lime and water are mixed and made into water content is 65%-70%(quality percentage composition), the medium of pH nature, wherein the mass ratio of apple branch wood chip, wheat bran, sucrose and white lime is that 80:18:1:1(is with dry weight basis).
In the above-mentioned cultivation method, described second class inoculum is with the female cultures that obtain 24 ℃-26 ℃ lower cultivations in described second class inoculum medium of planting of described light brown Aode mushroom (Oudemansiella canarii).
In the above-mentioned cultivation method, the female kind of institute's light brown Aode mushroom (Oudemansiella canarii) cultivated in mother culture media, and every liter of described mother culture media is made by following raw material: glucose 20g, soy peptone 4g, potato 200g, KH
2PO
41g, MgSO
40.5g, the water of vitamin C 10mg, agar 16g, surplus; The pH of described mother culture media is 8.5.
Another object of the present invention provides a kind of fruit body medium of light brown Aode mushroom.
The fruit body medium of light brown Aode mushroom provided by the present invention is that cotton seed hulls, apple branch wood chip, wheat bran, white lime and water are mixed and made into water content is 65%-70%(quality percentage composition) medium, wherein, the proportioning of cotton seed hulls, apple branch wood chip, wheat bran, white lime is 1)-5) in any:
1）50：30：18：2；
2）42：38：18：2；
3）（42-50）：（30-38）：18：2；
4）（50-65）：(15-30）：18：2；
5）42-65：15-38：18：2。
Wherein, the C/N ratio of described fruit body medium can be 50-60,56-60,50-56,56,60 or 50.
A further object of the present invention provides the culture medium for cultivating of light brown Aode mushroom.
The culture medium for cultivating of light brown Aode mushroom provided by the present invention comprises the fruit body medium of above-mentioned light brown Aode mushroom.
Further, the culture medium for cultivating of described light brown Aode mushroom also comprises the second class inoculum medium; Described second class inoculum medium is that apple branch wood chip, wheat bran, sucrose, white lime and water are mixed and made into water content is 65%-70%(quality percentage composition) medium, wherein the mass ratio of apple branch wood chip, wheat bran, sucrose and white lime is 80:18:1:1.
Further, the culture medium for cultivating of described light brown Aode mushroom also comprises mother culture media, and every liter of described mother culture media is made by following raw material: glucose 20g, soy peptone 4g, potato 200g, KH
2PO
41g, MgSO
40.5g, the water of vitamin C 10mg, agar 16g, surplus; The pH of described mother culture media is 8.5.
The biological efficiency of the cultivation method of light brown Aode mushroom of the present invention reaches 109%-123%, needs 31-42 days from bacterial classification being accessed the fruit body medium to the first damp mushroom of gathering, the 55-79 days altogether whole cultivation production cycle of the four damp mushrooms of gathering.
Description of drawings
Fig. 1 is the wild sporophore shape of light brown Aode mushroom (Oudemansiella canarii).
Fig. 2 is light brown Aode mushroom (Oudemansiella canarii) basidiospore form.
Bar length among the figure is 50.0 μ m.
Fig. 3 is light brown Aode mushroom (Oudemansiella canarii) artificial cultivation fruit body.
Embodiment
Following embodiment is convenient to understand better the present invention, but does not limit the present invention.Experimental technique among the following embodiment if no special instructions, is conventional method.
Used material, reagent etc. if no special instructions, all can obtain from commercial channels among the following embodiment.
Light brown Aode mushroom among the following embodiment (Oudemansiella canarii) (Yang Zhuliang, a surname is solemn. the classification of Southwestern China Genus Oudeman Siella Spec [J]. and fungi journal, 1993,12 (1): 16 ~ 27; [2] Li Jidong, Lin Yuexin. Oudemansiella canarii deep layer fermenting process research [J]. edible mushroom journal, 2003,10 (1): 46 ~ 51; Fourth of the twelve Earthly Branches morning mist. Macrofungi From China [M]. Zhengzhou: Henan science tech publishing house, 2000:113) pick up from Chinese yunnan, its bacteria cover diameter 4-10cm, first flat hemispherical, there is mucus on the surface; Initial stage central authorities' brown, edge be shallow (Figure 1A) gradually, and the later stage becomes greyish white to Slate grey; Bacterial context white, colloid; Stem is often crooked, long 5-16cm, and thick 5-15mm, dirty white, there are dark brown cilium and vertical stripe in the surface; Lamella white is to dirty white, and is rarer, not isometric, straight giving birth to prolonging life; Spore print white, the spore oval is extremely subsphaeroidal, diameter 14-16 μ m * 16-18 μ m(Figure 1B).The public can from field acquisition, also can obtain from the Beijing City Agriculture and Forestry Institute this bacterial classification.
Medium among the following embodiment is as follows:
1, comprehensive PDA culture medium
Made by following raw material for every liter: potato 200g, agar 20g, glucose 20g, soy peptone 5g, KH
2PO
43g, MgSO
41.5g, Cobastab
1The water of 10mg, surplus.121 ℃ of autoclavings 30 minutes.
2, basal medium
Made by following raw material for every liter: glucose 20g, soy peptone 2g, KH
2PO
41g, MgSO
40.5g, Cobastab
1The water of 10mg, agar 16g, surplus.121 ℃ of autoclavings 30 minutes.
3, enriched medium
Made by following raw material for every liter: glucose 20g, soy peptone 2g, potato 200g, KH
2PO
41g, MgSO
40.5g, Cobastab
1The water of 10mg, agar 16g, surplus.121 ℃ of autoclavings 30 minutes.
Adopt following assay method among the following embodiment:
(1) Growth rate is measured: light brown Aode mushroom (Oudemansiella canarii) bacterial classification inoculation is in the central authorities on plating medium surface, cultivate after 10 days, measure colony diameter, be the per day growth rate of mycelia with the bacterium colony mean radius divided by fate.
(2) the mycelia dry weight is measured: the plate of having measured growth rate, put into 121 ℃ of pressure cookers and be heated to medium thawing (10min), while hot light brown Aode mushroom (Oudemansiella canarii) mycelia is taken out and place the individual layer gauze, hot bath 1h, then place interior 105 ℃ of drying baker to dry to constant weight weighing.
(3) growth potential is measured: light brown Aode mushroom (Oudemansiella canarii) mycelial growth is dense, and growing way is vigorous, with " ++ ++ " expression; Mycelial growth is closeer, and growing way is better, with " +++" expression; Mycelial growth is close, and growing way is general, with " ++ " expression; Mycelial growth is rare, growing way a little less than, with "+" expression; Mycelia does not grow, with "-" expression.
(4) biological efficiency calculates: biological efficiency (%)=(bright mushroom weight/composts or fertilisers of cultivating dry weight) * 100%
Wherein, the assay method of medium or material dry weight is as follows: place in the drying baker 105 ℃ to dry to constant weight weighing medium or material.
Embodiment 1, cultivation light brown Aode mushroom (Oudemansiella canarii)
One, the female preparation of planting of light brown Aode mushroom (Oudemansiella canarii)
1, adopt tissue isolation to be inoculated in the inclined-plane that above-mentioned comprehensive PDA culture medium is housed from the cap isolate piece of light brown Aode mushroom (Oudemansiella canarii), cultivated 7-8 days for 25 ℃, grow the mycelia of light brown Aode mushroom (Oudemansiella canarii), obtain the female kind of light brown Aode mushroom (Oudemansiella canarii).
2, the optimization of mother culture media and condition of culture
(1) the synchronously preparation of mycelia: with the female kind of the light brown Aode mushroom (Oudemansiella canarii) that separates flat board (diameter 9cm) central authorities that picking one fritter strain transfer is made to comprehensive PDA culture medium in superclean bench, place 25 ℃ of incubator lucifuges to cultivate, treat to use when mycelia is covered with plate soon card punch (6mm) to beat the mycelia piece at the circle of dull and stereotyped same diameter, guarantee that the bacterium ferfas of subsequent experimental is identical age.
(2) carbon source experiment: the phosphorus content of 20g glucose is as standard in every liter of basal medium, usefulness and 20g glucose equate eight kinds of carbon sources of sucrose, maltose, lactose, starch, sorbierite, mannitol, sodium carboxymethylcellulose of phosphorus content respectively, replace glucose in the basal medium, every processing repeats for 5 times, the pH nature, lucifuge is cultivated in 25 ℃ of constant incubators, (is made by following raw material for every liter: soy peptone 2g, KH with carbonaceous sources medium not
2PO
41g, MgSO
40.5g, Cobastab
1The water of 10mg, agar 16g, surplus) do contrast (CK), observe different carbon sources on the impact of mycelial growth.
Experimental result is as shown in table 1, and there were significant differences for light brown Aode mushroom between different carbon sources (Oudemansiella canarii) mycelial growth rate, mycelium dry weight and growing way.Wherein, day growth speed on the medium take glucose as carbon source, mycelial growth potential is vigorous, reaches 5.45mm/d; Secondly be sucrose and maltose, day growth speed is respectively 4.81mm/d and 4.67mm/d; Being that day growth speed is the slowest on the medium of carbon source at lactose, is 2.01mm/d only, and much smaller than contrast, this may be because lactose has certain inhibitory action to this bacterial strain mycelia.Aspect the mycelia dry weight, the highest on the medium take glucose as carbon source, be 0.0530g; Sodium carboxymethylcellulose is minimum, only is 0.0094g; Wherein lactose and contrast are not remarkable for the mycelia dry weight difference of carbon source.Therefore, comprehensive above the analysis, the optimum carbon source that this experiment is chosen is glucose.
Table 1 carbon source is on the impact of light brown Aode mushroom mycelial growth
| Carbon source | Per day growth rate (mm/d) | Dry weight (g) | Mycelial growth potential |
| Glucose | 5.45±0.15a | 0.0530±0.0046a | ++++ |
| Sucrose | 4.81±0.13b | 0.0240±0.0007c | +++ |
| Maltose | 4.67±0.24b | 0.0291±0.0015b | +++ |
| Mannitol | 4.06±0.22c | 0.0270±0.0011bc | +++ |
| Starch | 3.88±0.11cd | 0.0261±0.0018bc | ++ |
| Sodium carboxymethylcellulose | 3.79±0.04d | 0.0094±0.0015e | ++ |
| Sorbierite | 3.69±0.02de | 0.0271±0.0023bc | ++ |
| CK | 3.50±0.04e | 0.0147±0.0002d | ++ |
| Lactose | 2.01±0.17f | 0.0131±0.0006d | + |
Annotate: numeral is mean+SD (n=5) in the table; Different English lower case represent 5% significance level (p＜0.05).
(3) nitrogenous source test: the nitrogen content of the 2g soy peptone in every liter of basal medium is as standard, usefulness and 2g soy peptone equate that the yeast of nitrogen content soaks powder, beef extract, ammonium nitrate, ammonium sulfate, urea, analysis for soybean powder, seven kinds of nitrogenous sources of glycine respectively, replace the peptone in the medium, the pH nature, lucifuge is cultivated in 25 ℃ of constant incubators, every processing 5 times repeats, and (is made by following raw material for every liter: glucose 20g, KH with nonnitrogenous source medium
2PO
41g, MgSO
40.5g, Cobastab
1The water of 10mg, agar 16g, surplus) do contrast (CK), the observation different nitrogen sources is on the impact of mycelial growth.
Experimental result is as shown in table 2, and light brown Aode mushroom (Oudemansiella canarii) has significant difference in different nitrogenous source medium growths.Mycelia day growth speed is the fastest with soy peptone and analysis for soybean powder, is respectively 5.63mm/d and 5.60mm/d, and the two difference is not remarkable; Next is beef extract; The slowest is contrast.From the mycelia dry weight, soy peptone is the highest, is 0.0542g; Secondly be analysis for soybean powder, it is the poorest to contrast.From the mycelia growing way, the mycelia take soy peptone as nitrogenous source is dense, and growing way is best; Secondly be analysis for soybean powder and beef extract; Urea and contrast are the poorest.Therefore, take the mycelia dry weight as main indexes, ligative hyphae growth rate and growth potential, this experiment selects soy peptone as optimum nitrogen source.
Table 2 nitrogenous source is on the impact of light brown Aode mushroom mycelial growth
| Nitrogenous source | Per day growth rate (mm/d) | Dry weight (g) | Mycelial growth potential |
| Soy peptone | 5.63±0.09a | 0.0542±0.0061a | ++++ |
| Analysis for soybean powder | 5.60±0.11a | 0.0383±0.0041b | +++ |
| Beef extract | 4.40±0.26b | 0.0348±0.0017bc | +++ |
| Urea | 3.92±0.06c | 0.0214±0.0010d | + |
| Yeast soaks powder | 3.59±0.13d | 0.0325±0.0011c | ++ |
| Ammonium nitrate | 3.43±0.26de | 0.0192±0.0028d | ++ |
| Ammonium sulfate | 3.29±0.10e | 0.0181±0.0006d | ++ |
| Glycine | 2.91±0.09f | 0.0172±0.0008d | ++ |
| CK | 2.78±0.04f | 0.0090±0.0021e | + |
Annotate: numeral is mean+SD (n=5) in the table; Different English lower case represent 5% significance level (p＜0.05).
(4) C/N ratio test: in every liter of basal medium, add the Different Weight peptone, being made into the medium C/N ratio is 10/1,20/1,30/1,40/1,50/1,60/1, the pH nature, lucifuge is cultivated in 25 ℃ of constant incubators, every processing repeats for 5 times, observes different C/N ratios on the impact of mycelial growth.
Experimental result is as shown in table 3, and different C/N ratios have a significant effect to light brown Aode mushroom (Oudemansiella canarii) mycelial growth.Light brown Aode mushroom (Oudemansiella canarii) mycelia all can grow in C/N ratio 10/1-60/1 scope, but the fastest with 20/1 C/N ratio mycelial growth rate, and compares significant difference with other C/N ratios.From the mycelia dry weight, mycelia is between 10/1-20/1, and the mycelia dry weight is the highest, and the two difference is not remarkable, but with other C/N ratio significant differences.Therefore, comprehensive above the analysis, it is 20/1 that C/N ratio is selected in this experiment.
Table 3 C/N ratio is on the impact of light brown Aode mushroom mycelial growth
| C/N ratio | Per day growth rate (mm/d) | Dry weight (g) | Mycelial growth potential |
| 20/1 | 6.46±0.21a | 0.0434±0.0045a | ++++ |
| 50/1 | 5.61±0.35b | 0.0232±0.0052b | ++ |
| 40/1 | 5.50±0.62bc | 0.0203±0.0025b | ++ |
| 30/1 | 5.34±0.30bc | 0.0267±0.0043b | ++ |
| 10/1 | 5.31±0.30bc | 0.0415±0.0053a | +++ |
| 60/1 | 4.85±0.32c | 0.0196±0.0016b | + |
Annotate: numeral is mean+SD (n=5) in the table; Different English lower case represent 5% significance level (p＜0.05).
(5) growth factor test: the Cobastab of the quality such as using respectively
2, Cobastab
6, vitamin C, corn steep liquor, inositol replace the Cobastab in the basal medium
1, (made by following raw material for every liter: glucose 20g, KH with the medium that do not add growth factor
2PO
41g, MgSO
40.5g, the water of agar 16g, surplus) do contrast (CK), observe different growth factors on the impact of light brown Aode mushroom (Oudemansiella canarii) mycelial growth.
Experimental result is as shown in table 4, and there is certain impact in different growth factors to light brown Aode mushroom (Oudemansiella canarii) strain growth speed, and wherein ascorbic mycelial growth rate is the fastest, is 5.35mm/d, but remarkable with other factor difference.From the mycelia dry weight, the mycelium dry weight of vitamin C and vitamin B1 is the highest, is respectively 0.0539g and 0.0509g, and is remarkable with other factor difference.From the mycelia growing way, growing way difference is little between each factor.Comprehensive above the analysis, this experiment selects vitamin C as the optimum growh factor.
Table 4 growth factor is on the impact of light brown Aode mushroom mycelial growth
| Growth factor | Per day growth rate (mm/d) | Dry weight (g) | Mycelial growth potential |
| Vitamin C | 5.35±0.09a | 0.0539±0.0042a | +++ |
| Corn steep liquor | 4.81±0.37b | 0.0371±0.0016c | +++ |
| Inositol | 4.68±0.31bc | 0.0442±0.0062b | ++ |
| Cobastab 6 | 4.64±0.04bc | 0.0449±0.0014b | ++ |
| Cobastab 1 | 4.57±0.16bcd | 0.0509±0.0033a | ++++ |
| Cobastab 2 | 4.31±0.21cd | 0.0376±0.0022c | ++++ |
| CK | 4.18±0.19d | 0.0347±0.0021c | + |
Annotate: numeral is mean+SD (n=5) in the table; Different English lower case represent 5% significance level (p＜0.05).
(6) temperature screening test: with carbon source, the nitrogenous source in the suitableeest carbon nitrogen source replacing of the above-mentioned test basal medium, adjusting C/N ratio is the suitableeest C/N ratio of above-mentioned test, select the most suitable growth factor, be made into and add rich Optimal Medium and (made by following raw material for every liter: glucose 20g, soy peptone 4g, potato 200g, KH
2PO
41g, MgSO
40.5g, the water of vitamin C 10mg, agar 16g, surplus), respectively 16,20,24,28,32,36 ℃ of lower constant temperature culture, the pH nature, every processing repeats for 5 times, the observation different temperatures is on the impact of mycelial growth.
Experimental result is as shown in table 5, and treatment of different temperature has significant impact to light brown Aode mushroom (Oudemansiella canarii) strain growth speed.Be between 24 ℃-32 ℃ in temperature, but the equal normal growth of mycelia; Wherein, mycelial growth is rapid in the time of 28 ℃, and day growth speed, mycelium dry weight, growth potential are optimum, and each is processed and compares significant difference with other.When temperature was lower than 24 ℃, along with the reduction of temperature, mycelial growth rate slowed down; When temperature reached 36 ℃, mycelia almost stopped growing.It is therefore, comprehensive above that to analyze the suitableeest cultivation temperature that this experiment chooses be 28 ℃.
Table 5 temperature is on the impact of light brown Aode mushroom mycelial growth
| Temperature | Per day growth rate (mm/d) | Dry weight (g) | Mycelial growth potential |
| 28℃ | 7.09±0.08a | 0.0473±0.0020a | ++++ |
| 24℃ | 6.11±0.10b | 0.0281±0.0017c | +++ |
| 32℃ | 5.66±0.08c | 0.0373±0.0011b | +++ |
| 20℃ | 4.39±0.09d | 0.0059±0.0019e | ++ |
| 16℃ | 2.09±0.08e | 0.0098±0.0005d | + |
| 36℃ | 1.14±0.05f | 0.0083±0.0008de | + |
Annotate: numeral is mean+SD (n=5) in the table; Different English lower case represent 5% significance level (p＜0.05).
(7) pH screening test: adopt to add rich Optimal Medium and (made by following raw material for every liter: glucose 20g, soy peptone 4g, potato 200g, KH
2PO
41g, MgSO
40.5g, the water of vitamin C 10mg, agar 16g, surplus), adjust pH is 5.0,5.5,6.0,6.5,7.0,7.5,8.0,8.5,9.0 respectively, and at 28 ℃, lucifuge is cultivated in the constant incubator, every processing repeats for 5 times, observes different pH on the impact of mycelial growth.
Experimental result is as shown in table 6, and light brown Aode mushroom (Oudemansiella canarii) mycelia is that 5.0-9.0 can both grow at pH, along with the rising of pH, growth rate has gradually and accelerates, growth potential is grow gradually also, but when pH was 9.0, mycelial growth rate, growth potential descended.Be 8.5 o'clock at pH, mycelial growth is the fastest, and mycelia is dense, and day growth speed is 7.76mm/d, and processes significant difference with other; Secondly be pH7.5, pH9.0 and pH8.0; PH is that 5.0 o'clock mycelial growth rates are the slowest.Aspect the mycelia dry weight, pH is that 8.5 o'clock mycelia dry weights are the highest, is 0.0549g, with each processes that to compare difference not remarkable under other alkali conditions.Therefore, take the mycelia dry weight as main indexes, ligative hyphae growth rate and growth potential, the suitable growing environment of light brown Aode mushroom mycelia is alkalescence, optimal pH is 8.5.
Table 6pH is on the impact of light brown Aode mushroom mycelial growth
| pH | Per day growth rate (mm/d) | Dry weight (g) | Mycelial growth potential |
| 8.5 | 7.76±0.23a | 0.0549±0.0065a | ++++ |
| 7.5 | 7.15±0.08b | 0.0510±0.0095ab | +++ |
| 9.0 | 6.97±0.15bc | 0.0436±0.0139abc | ++ |
| 8.0 | 6.79±0.37bcd | 0.0403±0.0036abc | ++ |
| 5.5 | 6.52±0.21cde | 0.0371±0.0062bc | + |
| 7.0 | 6.49±0.21de | 0.0465±0.0103abc | ++ |
| 6.0 | 6.47±0.53de | 0.0454±0.0027abc | ++ |
| 6.5 | 6.20±0.10e | 0.0517±0.0067ab | ++ |
| 5.0 | 6.14±0.12e | 0.0357±0.0022c | + |
Annotate: numeral is mean+SD (n=5) in the table; Different English lower case represent 5% significance level (p＜0.05).
To sum up, the female optimal culture condition of planting of light brown Aode mushroom (Oudemansiella canarii) is that to adopt pH be that 8.5 the rich Optimal Medium that adds (is made by following raw material for every liter: glucose 20g, soy peptone 4g, potato 200g, KH
2PO
41g, MgSO
40.5g, the water of vitamin C 10mg, agar 16g, surplus), 28 ℃ of lower constant temperature culture.
Two, the preparation of secondary kind
It is 65%-70%(quality percentage composition that apple branch wood chip, wheat bran, sucrose, white lime and water are mixed and made into water content), the secondary kind medium of pH nature, wherein the mass ratio of apple branch wood chip, wheat bran, sucrose and white lime is that 80:18:1:1(is with dry weight basis).Secondary kind medium is sub-packed in the vial 121 ℃ of autoclavings 120 minutes.Wherein, the carbon nitrogen content in wheat bran and the apple branch wood chip is as shown in table 8.
Plant access and add rich Optimal Medium and (made by following raw material for every liter: potato 200g, glucose 20g, soy peptone 4g, KH the light brown Aode mushroom (Oudemansiella canarii) of step 1 is female
2PO
41g, MgSO
40.5g, the water of vitamin C 10mg agar powder 16g, surplus, pH 8.5) at 28 ℃ of lower constant temperature culture 7-10 days, be transferred in the above-mentioned secondary kind medium, cultivated 14-16 days 24 ℃-26 ℃ lower lucifuges, mycelia is covered with medium, obtains second class inoculum.
Three, inoculated and cultured
1, medium is made and pack
This experiment is adopted by three kinds of fruit body medium T1, T2 of cotton seed hulls, wheat bran, apple branch wood chip and white lime preparation and T3, and it forms table 7.Wherein " % " is the quality percentage composition.
Composition and the C/N ratio of table 7 fruit body medium
Carbon nitrogen content in cotton seed hulls in the above-mentioned medium, wheat bran and the apple branch wood chip is as shown in table 8.
The carbon nitrogen content of table 8 culture medium raw material
| ? | Full carbon (%) | Full nitrogen (%) | C:N |
| Cotton seed hulls | 38.06 | 0.57 | 66.77 |
| Wheat bran | 36.92 | 1.98 | 18.65 |
| The apple branch wood chip | 41.34 | 0.12 | 358.61 |
Wherein, the mensuration of full carbon adopts potassium dichromate method, and full nitrogen determination adopts Kjeldahls method.Wherein " % " is the quality percentage composition.
The compound method of above-mentioned three kinds of fruit body medium is as follows: it is 65%-70%(quality percentage composition that cotton seed hulls, apple branch wood chip, wheat bran, white lime and water are mixed and made into water content), the medium of pH nature.Wherein among the fruit body medium T1, the mass ratio of cotton seed hulls, apple branch wood chip, wheat bran, white lime is that 65:15:18:2(is with dry weight basis); Among the fruit body medium T2, the mass ratio of cotton seed hulls, apple branch wood chip, wheat bran, white lime is that 50:30:18:2(is with dry weight basis); Among the fruit body medium T3, the mass ratio of cotton seed hulls, apple branch wood chip, wheat bran, white lime is that 42:38:18:2(is with dry weight basis).Medium is sub-packed in the high pressure resistant plastic sack of 17cm * 33cm * 0.04cm, every packed siccative (other outer components dewater in the fruit body medium) 380g.121 ℃ of autoclavings 2 hours obtain being equipped with the bacterium bag of fruit body medium.
2, inoculation and a bacterium
Light brown Aode mushroom (Oudemansiella canarii) second class inoculum of step 2 preparation is evenly accessed the bacterium bag that step 1 is equipped with above-mentioned fruit body medium, and every bag of inoculum concentration is 3.14 * 10
-5m
3Experiment arranges three repetitions, and each repeats to cultivate 100 bacterium bags, every bag culture medium dry weight 380g.
Inoculation is rear to be 24 ℃-26 ℃ in environmental temperature, and relative air humidity is 50%-60%, the well-ventilated, and lucifuge is cultivated, and mycelia is covered with medium after 21-23 days.
3, low temperature stimulation
The bacterium bag is transferred under-5 ℃ of-5 ℃ of conditions processed 3-5 days, finish low temperature stimulation;
4, fruiting
It is 23 ℃-26 ℃ that the bacterium bag is transferred to environmental temperature, relative air humidity 80%-90%, intensity of illumination 300Lux, the well-ventilated, gas concentration lwevel 1500ppm(ppm: 1,000,000 air molecules have a carbon dioxide molecule) condition under cultivate and formed to fruit body primordium in 5-7 days;
5, gather
Continue to cultivate after 3-5 days, the fruit body stem grows to about 10cm, and cap launches, but spore is when not yet launching, the first damp mushroom (Fig. 3) of gathering.Stop water spray after the harvesting, the maintenance relative air humidity is 50%-60%, the well-ventilated, mycelia was recovered 5-7 days, then be 23 ℃-26 ℃ according to step 4 fruiting condition in environmental temperature, relative air humidity 80%-90%, intensity of illumination 300Lux, the well-ventilated, gas concentration lwevel 1500ppm(ppm: 1,000,000 air molecules have a carbon dioxide molecule) condition under cultivated 3-5 days, the second damp mushroom of gathering, after plucking, the second damp mushroom treats that mycelia recovered 5-7 days, be 23 ℃-26 ℃ according to step 4 fruiting condition in environmental temperature, relative air humidity 80%-90%, intensity of illumination 300Lux, the well-ventilated, gas concentration lwevel 1500ppm(ppm: 1,000,000 air molecules have a carbon dioxide molecule) condition under cultivated 3-5 days, the 3rd damp mushroom of gathering, treating after the 3rd damp mushroom is plucked that mycelia recovered 5-7 days, is 23 ℃-26 ℃ according to step 4 fruiting condition in environmental temperature, relative air humidity 80%-90%, intensity of illumination 300Lux, the well-ventilated, gas concentration lwevel 1500ppm(ppm: 1,000,000 air molecules have a carbon dioxide molecule) condition under cultivated the 4th damp mushroom of gathering 3-5 days.
The result shows, during fruit body medium T1 processes, the bacteria developing period of light brown Aode mushroom (Oudemansiella canarii) needs 20-22 days, low temperature stimulation 3-5 days, fruit body primordium forms phase 5-8 days under the fruiting condition, the fruit body differentiation is to gathering 3-5 days, and calculating needs 31-40 days from being seeded to the first damp mushroom of gathering.Mushroom tide interval 5-8 days, every damp massee fruiting bodies differentiation is to gathering 3-5 days; Gather 55-79 days altogether whole cultivation production cycles of four damp mushrooms.
During fruit body medium T2 processes, the bacteria developing period of light brown Aode mushroom (Oudemansiella canarii) needs 21-23 days, low temperature stimulation 3-5 days, fruit body primordium forms phase 5-7 days under the fruiting condition, the fruit body differentiation is to gathering 3-5 days, and calculating needs 32-40 days from being seeded to the first damp mushroom of gathering.Mushroom tide interval 5-7 days, every damp massee fruiting bodies differentiation is to gathering 3-5 days; Gather 56-76 days altogether whole cultivation production cycles of four damp mushrooms.
During fruit body medium T3 processes, the bacteria developing period of light brown Aode mushroom (Oudemansiella canarii) needs 22-25 days, low temperature stimulation 3-5 days, fruit body primordium forms phase 5-7 days under the fruiting condition, the fruit body differentiation is to gathering 3-5 days, and calculating needs 33-42 days from being seeded to the first damp mushroom of gathering.Mushroom tide interval 5-7 days, every damp massee fruiting bodies differentiation is to gathering 3-5 days; Gather 57-78 days altogether whole cultivation production cycles of four damp mushrooms.
With " biological efficiency " expression output.Described biological efficiency refers to the ratio of edible mushroom fresh weight and used composts or fertilisers of cultivating dry weight (other outer components dewater in the fruit body medium), and percentage commonly used represents.Produced 80kg fresh food bacterium such as the dried composts or fertilisers of cultivating of 100kg, then the biological efficiency of this edible mushroom is 80%.
Under this experiment cultivation condition, the biological efficiency of light brown Aode mushroom (Oudemansiella canarii) on the fruit body medium is as shown in table 9, the biological efficiency at fruit body medium treatment T2 the highest (calculating by four damp mushroom output) wherein, be 122.66% ± 5.11%, document (Marcelo J.S.R.
M.T.T., Vera L.R.B., Marina C..Cultivation of the edible mushroom Oudemansiella canarii (Jungh.)
Inlignocellulosic substrates.Brazilian Journal of Microbiology, 2001, in 32:211-214.) take as prescription biological efficiency (50.66% ± 20.41%) 2.42 times of bagasse.
Output (biological efficiency) statistics (unit: %) of table 9 light brown Aode mushroom
Annotate: in the 5% horizontal significance, have between the processing of same letter without significant difference, there were significant differences between alphabetical diverse processing.
Claims (9)
1. the cultivation method of light brown Aode mushroom, comprise the step with fruit body medium culture light brown Aode mushroom, described fruit body medium is that cotton seed hulls, apple branch wood chip, wheat bran, white lime and water are mixed and made into water content is 65%-70%(quality percentage composition) medium, wherein the proportioning of cotton seed hulls, apple branch wood chip, wheat bran, white lime is 1)-5) in any:
1）50：30：18：2；
2）42：38：18：2；
3）（42-50）：（30-38）：18：2；
4）（50-65）：（15-30）：18：2；
5）42-65：15-38：18：2。
2. cultivation method according to claim 1, it is characterized in that: described cultivation method comprises the steps:
1) send out bacterium: light brown Aode mushroom (Oudemansiella canarii) second class inoculum is accessed described fruit body medium, is 24 ℃-26 ℃ in environmental temperature, and relative air humidity is to be cultured to mycelia under the condition of 50%-60% to cover with medium;
2) low temperature stimulation: the culture of step 1) is transferred under-5 ℃ of-5 ℃ of conditions processed 3-5 days, finish low temperature stimulation;
3) fruiting; It is 23 ℃-26 ℃ that the culture of step 1) is transferred to environmental temperature, and relative air humidity 80%-90%, intensity of illumination 300-600Lux, gas concentration lwevel are cultured to fruit body primordium under the condition of 1500-2000ppm to form.
3. cultivation method according to claim 2 is characterized in that: described second class inoculum is that female kind of light brown Aode mushroom (Oudemansiella canarii) cultivated the culture that obtains in the second class inoculum medium; Described second class inoculum medium is that apple branch wood chip, wheat bran, sucrose, white lime and water are mixed and made into water content is 65%-70%(quality percentage composition) medium, wherein the mass ratio of apple branch wood chip, wheat bran, sucrose and white lime is 80:18:1:1.
4. cultivation method according to claim 3 is characterized in that: described second class inoculum is to plant in described second class inoculum medium at 24 ℃-26 ℃ lower cultures that obtain of cultivating described light brown Aode mushroom (Oudemansiella canarii) is female.
5. according to claim 3 or 4 described cultivation methods, it is characterized in that: the female kind of institute's light brown Aode mushroom (Oudemansiella canarii) cultivated in mother culture media, and every liter of described mother culture media is made by following raw material: glucose 20g, soy peptone 4g, potato 200g, KH
2PO
41g, MgSO
40.5g, the water of vitamin C 10mg, agar 16g, surplus; The pH of described mother culture media is 8.5.
6. the fruit body medium of light brown Aode mushroom (Oudemansiella canarii), described fruit body medium is that cotton seed hulls, apple branch wood chip, wheat bran, white lime and water are mixed and made into water content is 65%-70%(quality percentage composition) medium, wherein the proportioning of cotton seed hulls, apple branch wood chip, wheat bran, white lime is 1)-5) in any:
1）50：30：18：2；
2）42：38：18：2；
3）（42-50）：（30-38）：18：2；
4）（50-65）：（15-30）：18：2；
5）42-65：15-38：18：2。
7. the culture medium for cultivating of light brown Aode mushroom (Oudemansiella canarii) comprises the fruit body medium of claim 6.
8. medium according to claim 7, it is characterized in that: described culture medium for cultivating comprises the second class inoculum medium; Described second class inoculum medium is that apple branch wood chip, wheat bran, sucrose, white lime and water are mixed and made into water content is 65%-70%(quality percentage composition) medium, wherein the mass ratio of apple branch wood chip, wheat bran, sucrose and white lime is 80:18:1:1.
9. according to claim 7 or 8 described medium, it is characterized in that: described culture medium for cultivating comprises mother culture media, and every liter of described mother culture media is made by following raw material: glucose 20g, soy peptone 4g, potato 200g, KH
2PO
41g, MgSO
40.5g, the water of vitamin C 10mg, agar 16g, surplus; The pH of described mother culture media is 8.5.
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