CN105981636A - Peat moss protonema rapid reproduction method - Google Patents
Peat moss protonema rapid reproduction method Download PDFInfo
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- CN105981636A CN105981636A CN201510070439.6A CN201510070439A CN105981636A CN 105981636 A CN105981636 A CN 105981636A CN 201510070439 A CN201510070439 A CN 201510070439A CN 105981636 A CN105981636 A CN 105981636A
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Abstract
The invention relates to the field of organic synthesis, and discloses a peat moss protonema rapid reproduction method. The peat moss protonema rapid reproduction method includes the following steps: preparation of a culture medium, preparation of suspension liquid, cultivation, and continuous proliferation cultivation for generations. According to the method, reproduction of the peat moss can be performed at any time and cannot be affected by external factors such as seasons and environments; the method is low in cost, is less in occupation space, and can provide necessary growth elements and a growth environment for peat moss protonemata; the method can achieve rapid reproduction of the peat moss protonemata and can solve the problem that the growth speed is slow and the reproduction of the peat moss protonemata is greatly affected by the environment; a selected pH value is identical to that of meta-acid soil of a natural growth environment of the peat moss; cane sugar is added in the culture medium and provides a carbon source, and then the yield of the peat moss protonemata is greatly improved; the method selects 6-benzylamino adenine (6-BA) as cytokinin, which has a role greater than that of other auxins in the peat moss protonemata; the optimum concentration of the 6-BA is 0.05-0.2 [mu]mol/L, and the growth can be suppressed when the concentration of the 6-BA is greater than or less than the optimum concentration.
Description
Technical field
The present invention relates to horticultural gardening technical field, particularly relate to a kind of bog moss protonema rapid propagation method.
Background technology
Peat, can be directly as cultivation medium, it is possible to as the fertilizer of conditioned soil earth, in recent years rich in organic and humic acid, and the high speed development of horticulture industry has driven the formation of peat industry.But, the market demand of peat is on the increase, and peat wetland is the most constantly reduced by exhaustive exploitation, area, and China is gradually converted into importer from peat exported country.Vegetation distribution, ecological diversity are not only caused serious threat by the destruction of peat wetland, also result in a series of consequences such as the reduction of nature ability to pool carbon, human residential environment's deterioration.For realizing the sustainable use of mud and charcoal, not affecting the development of agricultural and horticulture industry, the succedaneum finding peat is problem demanding prompt solution simultaneously.
Bog moss (Sphagnum.sp.), also known as sphagna, is the general designation of Genus Sphagnum plant, is also the main source of formtion of peat.The American-European gardening developed country such as Germany has studied and bog moss has substituted peat as growth substrate, and is successfully applied to the cultivation of plants such as Cuculus polioephalus, brier.Therefore, the exploitation of bog moss and application can promote the development of horticulture industry.
Summary of the invention
The present invention is directed to existing in shortcoming, disclose a kind of bog moss protonema rapid propagation method.
In order to solve above-mentioned technical problem, the present invention is addressed by following technical proposals.
A kind of bog moss protonema rapid propagation method, comprises the following steps,
The most every 1 liter of water adds the ammonium nitrate of 0.1-0.3g, the calcium chloride of 0.05-0.15g, the potassium dihydrogen phosphate of 0.05-0.15g, the magnesium sulfate of 0.05-0.15g, the iron chloride of 0.02-0.035g；PH value is adjusted to 5-6, then adds the sucrose of 5-10g and the 6-benzyl aminoadenine of 0.05-0.2 μm ol, obtain culture medium；Autoclaving processes culture medium；
B. pulverize bog moss protonema, filter, the bog moss protonema crushed is moved in sterilized water, makes suspension；
C. adding suspension prepared by step B in culture medium prepared by step A, wherein add 0.5-1.5ml suspension in every 10ml culture medium, put into shaking table, rotating speed 130-150r/min, illumination is 2800-3500lux, and the photoperiod is day night 10 h/ 14 h；This condition of culture solve the bog moss protonema speed of growth slowly, the technical problem the most affected by environment.
D., after cultivating 20 days, bog moss protonema is again pulverized, in the culture medium that after filtration prepared by subpackage to step A, wherein every 10ml culture medium adds 0.5-1.5ml suspension, put into shaking table, rotating speed 130-150r/min, illumination is 2800-3500lux, and the photoperiod is day night 10 h/ 14 h；After cultivating 20 days, culture medium nutrient depletion in former bottle, the protonema of propagation is pulverized again.This condition of culture solves the technical problem that the bog moss protonema speed of growth is slow, reaches the purpose of fast breeding.
E. repeat step D, cultivate propagation number generation continuously.
The culture medium that the present invention uses is Beneck culture medium.The growth that culture medium is bog moss protonema provides necessary growth elements and environment, so the screening of culture medium occupies critical role during whole Fast-propagation.In research process, having carried out the screening of minimal medium, test medium is respectively as follows: Beneck, Knop, BCD, MS, 1/2MS, water.Being found by the culture experiment of different culture media, Beneck and the Knop culture medium of Low-salinity is relatively suitable for the growth of bog moss；It is that lichen is commonly used the BCD culture medium of culture medium and is then not suitable for bog moss growth equally；The variable concentrations MS of high salinity is less useful for the growth of bog moss, and even along with the prolongation of incubation time, protonema can gradually turn yellow.
In research process, having carried out the screening of pH value, test pH gradient is respectively 4,5,6,7,8.Result of study shows, bog moss protonema is also bred very fast in the culture medium that pH is relatively low, and pH5.0-6.0 is its optimum growth acid-base value, and along with the rising of pH, bog moss protonema increment reduces, even stops growing.This coincide with bog moss natural habitat meta-acid soil.
The sucrose added provides carbon source, greatly improves bog moss protonema yield.In research process, having carried out the screening of carbon source and concentration thereof, test carbon source is respectively sucrose and glucose, and concentration is respectively 5 g/L, 10 g/L, 20 g/L, 40g/L.Result of study shows, the interpolation of carbon source greatly improves bog moss protonema yield: sucrose and glucose all have remarkable result to it, but higher to the utilization ratio of sucrose, the sucrose (5-10g/L) of low concentration is more beneficial for the propagation of bog moss, and propagation efficiency processes many 2.7-4.3 times than high concentration.
The present invention selects 6-benzyl aminoadenine (6-BA) as the basic element of cell division, effect to bog moss protonema is more than other auxin, protonema is bred without remarkable effect by such as 6-BA by the effect of bog moss protonema more than auxin indolebutyric acid (IBA), IBA.It is 0.05-0.2 μm ol/L that the present invention filters out the optium concentration of hormone 6-BA, more than or less than this concentration Developing restraint on the contrary.In research process, having carried out the screening of hormone and concentration thereof, test hormone is respectively IBA and 6-BA, and concentration is respectively 0.1 μm ol/L, 1 μm ol/L, 10 μm ol/L.Result of study shows, basic element of cell division 6-BA is more than auxin IBA to the effect of bog moss protonema.IBA is less to bog moss protonema proliferation function, and high concentration (10 μm ol/L) also can suppress it to grow.Low concentration (0.05-0.2 μm ol/L) 6-BA significantly improves bog moss yield, weakens along with concentration raises facilitation.
As preferably, in step A, every 1 liter of water adds the ammonium nitrate of 0.2g, the calcium chloride of 0.1g, the potassium dihydrogen phosphate of 0.1g, the magnesium sulfate of 0.1g, the iron chloride of 0.028g；PH value is adjusted to 5-6, then adds the sucrose of 10g and the 6-benzyl aminoadenine of 0.1 μm ol, obtain culture medium.The formula of the optimal medium that the present invention filters out: add the ammonium nitrate of 0.2g, the calcium chloride of 0.1g, the potassium dihydrogen phosphate of 0.1g, the magnesium sulfate of 0.1g, the iron chloride of 0.028g in every 1 liter of water.The growth that culture medium is bog moss protonema provides necessary growth elements and environment, and the culture medium of this proportioning can Fast-propagation bog moss protonema.
As preferably, in step A, magnesium sulfate is MgSO4·7H2O。
As preferably, in step A, iron chloride is FeCl3
·6 H2O。
As preferably, in step A, culture medium being loaded in 500ml triangular flask, the most bottled about 200ml, autoclaving is stand-by.
As preferably, in step B, bog moss protonema pulverizes 1min at superclean bench, and specific breakage rate is 18000r/min, and 200 mesh cells are sieved through filter；The bog moss protonema crushed is moved in sterilized water, makes suspension.
As preferably, in step C, every 10ml culture medium adds 1ml suspension.
As preferably, in step C, putting into shaking table, rotating speed 140r/min, illumination is 3000lux, and the photoperiod is day night 10 h/ 14 h.
As preferably, in step D, putting into shaking table, rotating speed 140r/min, illumination is 3000lux, and the photoperiod is day night 10 h/ 14 h.
As preferably, in step D, every 10ml culture medium adds 1ml suspension.
Compared with prior art, the invention have the benefit that
The present invention cultivates and can breed at any time, is not affected by the external factor such as season, environment, and cost is relatively low, and the resource that takes up room is less；Growth for bog moss protonema provides necessary growth elements and environment.Meanwhile, it is capable to Fast-propagation bog moss protonema, solve the speed of growth big technical problem slow, affected by environment.
The optimal pH that the present invention selects is 5-6, coincide with bog moss natural habitat meta-acid soil.The sucrose that the present invention adds provides carbon source, greatly improves bog moss protonema yield.The optimum carbon source that the present invention selects is sucrose, and utilization ratio is the highest, and the yield of bog moss protonema reaches the highest.The present invention selects 6-benzyl aminoadenine (6-BA) as the basic element of cell division, effect to bog moss protonema is more than other auxin, protonema is bred without remarkable effect by such as 6-BA by the effect of bog moss protonema more than auxin indolebutyric acid (IBA), IBA.It is 0.05-0.2 μm ol/L that the present invention filters out the optium concentration of hormone 6-BA, more than or less than this concentration Developing restraint on the contrary.
Detailed description of the invention
Embodiment
1
A kind of bog moss protonema rapid propagation method, comprises the following steps,
The most every 1 liter of water adds the ammonium nitrate of 0.2g, the calcium chloride of 0.1g, the potassium dihydrogen phosphate of 0.1g, the magnesium sulfate of 0.1g, the iron chloride of 0.028g；PH value is adjusted to 5, then adds the sucrose of 10g and the 6-benzyl aminoadenine of 0.1 μm ol, obtain culture medium；Autoclaving processes culture medium；
B. pulverize bog moss protonema, filter, the bog moss protonema crushed is moved in sterilized water, makes suspension；
C. adding suspension prepared by step B in culture medium prepared by step A, wherein add 1ml suspension in every 10ml culture medium, put into shaking table, rotating speed 140r/min, illumination is 3000lux, and the photoperiod is day night 10 h/ 14 h；
D. after cultivating 20 days, again pulverizing bog moss protonema, in the culture medium that after filtration prepared by subpackage to step A, wherein add 1ml suspension in every 10ml culture medium, put into shaking table, rotating speed 140r/min, illumination is 3000lux, and the photoperiod is day night 10 h/ 14 h；
E. repeat step D, cultivate propagation number generation continuously.
Embodiment
2
A kind of bog moss protonema rapid propagation method, comprises the following steps,
The most every 1 liter of water adds the ammonium nitrate of 0.1g, the calcium chloride of 0.05g, the potassium dihydrogen phosphate of 0.05g, the magnesium sulfate of 0.05g, the iron chloride of 0.02g；PH value is adjusted to 5, then adds the sucrose of 5g and the 6-benzyl aminoadenine of 0.05 μm ol, obtain culture medium；Autoclaving processes culture medium；
B. bog moss protonema pulverizes 1min at superclean bench, and specific breakage rate is 18000r/min, and 200 mesh cells are sieved through filter；The bog moss protonema crushed is moved in sterilized water, makes suspension；
C. adding suspension prepared by step B in culture medium prepared by step A, wherein add 0.5ml suspension in every 10ml culture medium, put into shaking table, rotating speed 130r/min, illumination is 2800lux, and the photoperiod is day night 10 h/ 14 h；
D., after cultivating 20 days, bog moss protonema is again pulverized, in the culture medium that after filtration prepared by subpackage to step A, wherein every 10ml culture medium adds 0.5ml suspension, put into shaking table, rotating speed 130r/min, illumination is 2800lux, and the photoperiod is day night 10 h/ 14 h；
E. repeat step D, cultivate propagation number generation continuously.
Embodiment
3
A kind of bog moss protonema rapid propagation method, comprises the following steps,
The most every 1 liter of water adds the ammonium nitrate of 0.3g, the calcium chloride of 0.15g, the potassium dihydrogen phosphate of 0.15g, the magnesium sulfate of 0.15g, the iron chloride of 0.035g；PH value is adjusted to 6, then adds the sucrose of 10g and the 6-benzyl aminoadenine of 0.2 μm ol, obtain culture medium；Culture medium being loaded in 500ml triangular flask, the most bottled about 200ml, autoclaving is stand-by.
B. pulverize bog moss protonema, filter, the bog moss protonema crushed is moved in sterilized water, makes suspension；
C. adding suspension prepared by step B in culture medium prepared by step A, wherein add 1.5ml suspension in every 10ml culture medium, put into shaking table, rotating speed 150r/min, illumination is 3500lux, and the photoperiod is day night 10 h/ 14 h；
D., after cultivating 20 days, bog moss protonema is again pulverized, in the culture medium that after filtration prepared by subpackage to step A, wherein every 10ml culture medium adds 1.5ml suspension, put into shaking table, rotating speed 150r/min, illumination is 3500lux, and the photoperiod is day night 10 h/ 14 h；
E. repeat step D, cultivate propagation number generation continuously.
Embodiment
4
A kind of bog moss protonema rapid propagation method, comprises the following steps,
The most every 1 liter of water adds the ammonium nitrate of 0.15g, the calcium chloride of 0.08g, the potassium dihydrogen phosphate of 0.08g, the MgSO of 0.08g4·7H2The FeCl of O, 0.025g3 ·6 H2O；PH value is adjusted to 5.5, then adds the sucrose of 6g and the 6-benzyl aminoadenine of 0.08 μm ol, obtain culture medium；Autoclaving processes culture medium；
B. pulverize bog moss protonema, filter, the bog moss protonema crushed is moved in sterilized water, makes suspension；
C. adding suspension prepared by step B in culture medium prepared by step A, wherein add 0.8ml suspension in every 10ml culture medium, put into shaking table, rotating speed 135r/min, illumination is 2900lux, and the photoperiod is day night 10 h/ 14 h；
D., after cultivating 20 days, bog moss protonema is again pulverized, in the culture medium that after filtration prepared by subpackage to step A, wherein every 10ml culture medium adds 0.8ml suspension, put into shaking table, rotating speed 135r/min, illumination is 2900lux, and the photoperiod is day night 10 h/ 14 h；
E. repeat step D, cultivate propagation number generation continuously.
Embodiment
5
A kind of bog moss protonema rapid propagation method, comprises the following steps,
The most every 1 liter of water adds the ammonium nitrate of 0.28g, the calcium chloride of 0.12g, the potassium dihydrogen phosphate of 0.12g, the magnesium sulfate of 0.14g, the iron chloride of 0.032g；PH value is adjusted to 6, then adds the sucrose of 9g and the 6-benzyl aminoadenine of 0.18 μm ol, obtain culture medium；Autoclaving processes culture medium；
B. pulverize bog moss protonema, filter, the bog moss protonema crushed is moved in sterilized water, makes suspension；
C. adding suspension prepared by step B in culture medium prepared by step A, wherein add 1.2ml suspension in every 10ml culture medium, put into shaking table, rotating speed 140r/min, illumination is 3400lux, and the photoperiod is day night 10 h/ 14 h；
D., after cultivating 20 days, bog moss protonema is again pulverized, in the culture medium that after filtration prepared by subpackage to step A, wherein every 10ml culture medium adds 1.2ml suspension, put into shaking table, rotating speed 140r/min, illumination is 3400lux, and the photoperiod is day night 10 h/ 14 h；
E. repeat step D, cultivate propagation number generation continuously.
Embodiment
6
A kind of bog moss protonema rapid propagation method, comprises the following steps,
The most every 1 liter of water adds the ammonium nitrate of 0.2g, the calcium chloride of 0.1g, the potassium dihydrogen phosphate of 0.1g, the magnesium sulfate of 0.1g, the iron chloride of 0.028g；PH value is adjusted to 5, then adds the sucrose of 10g and the 6-benzyl aminoadenine of 0.1 μm ol, obtain culture medium；Autoclaving processes culture medium；
B. bog moss protonema pulverizes 1min at superclean bench, and specific breakage rate is 18000r/min, and 200 mesh cells are sieved through filter；The bog moss protonema crushed is moved in sterilized water, makes suspension；
C. adding suspension prepared by step B in culture medium prepared by step A, wherein add 1ml suspension in every 10ml culture medium, put into shaking table, rotating speed 140r/min, illumination is 3000lux, and the photoperiod is day night 10 h/ 14 h；
D. after cultivating 20 days, again pulverizing bog moss protonema, in the culture medium that after filtration prepared by subpackage to step A, wherein add 1ml suspension in every 10ml culture medium, put into shaking table, rotating speed 140r/min, illumination is 3000lux, and the photoperiod is day night 10 h/ 14 h；
E. repeat step D, cultivate propagation number generation continuously.
Embodiment
7
A kind of bog moss protonema rapid propagation method, comprises the following steps,
The most every 1 liter of water adds the ammonium nitrate of 0.2g, the calcium chloride of 0.1g, the potassium dihydrogen phosphate of 0.1g, the MgSO of 0.1g4·7H2The FeCl of O, 0.028g3 ·6 H2O；PH value is adjusted to 5, then adds the sucrose of 10g and the 6-benzyl aminoadenine of 0.1 μm ol, obtain culture medium；Autoclaving processes culture medium；
B. pulverize bog moss protonema, filter, the bog moss protonema crushed is moved in sterilized water, makes suspension；
C. adding suspension prepared by step B in culture medium prepared by step A, wherein add 1ml suspension in every 10ml culture medium, put into shaking table, rotating speed 140r/min, illumination is 3000lux, and the photoperiod is day night 10 h/ 14 h；
D. after cultivating 20 days, again pulverizing bog moss protonema, in the culture medium that after filtration prepared by subpackage to step A, wherein add 1ml suspension in every 10ml culture medium, put into shaking table, rotating speed 140r/min, illumination is 3000lux, and the photoperiod is day night 10 h/ 14 h；
E. repeat step D, cultivate propagation number generation continuously.
In a word, the foregoing is only presently preferred embodiments of the present invention, all impartial changes made according to scope of the present invention patent and modification, all should belong to the covering scope of patent of the present invention.
Claims (10)
1. a bog moss protonema rapid propagation method, it is characterised in that: comprise the following steps,
The most every 1 liter of water adds the ammonium nitrate of 0.1-0.3g, the calcium chloride of 0.05-0.15g, the potassium dihydrogen phosphate of 0.05-0.15g, the magnesium sulfate of 0.05-0.15g, the iron chloride of 0.02-0.035g；PH value is adjusted to 5-6, then adds the sucrose of 5-10g and the 6-benzyl aminoadenine of 0.05-0.2 μm ol, obtain culture medium；Autoclaving processes culture medium；
B. pulverize bog moss protonema, filter, the bog moss protonema crushed is moved in sterilized water, makes suspension；
C. adding suspension prepared by step B in culture medium prepared by step A, wherein add 0.5-1.5ml suspension in every 10ml culture medium, put into shaking table, rotating speed 130-150r/min, illumination is 2800-3500lux, and the photoperiod is day night 10 h/ 14 h；
D., after cultivating 20 days, bog moss protonema is again pulverized, in the culture medium that after filtration prepared by subpackage to step A, wherein every 10ml culture medium adds 0.5-1.5ml suspension, put into shaking table, rotating speed 130-150r/min, illumination is 2800-3500lux, and the photoperiod is day night 10 h/ 14 h；
E. repeat step D, cultivate propagation number generation continuously.
Bog moss protonema rapid propagation method the most according to claim 1, it is characterised in that: in step A, every 1 liter of water adds the ammonium nitrate of 0.2g, the calcium chloride of 0.1g, the potassium dihydrogen phosphate of 0.1g, the magnesium sulfate of 0.1g, the iron chloride of 0.028g；PH value is adjusted to 5-6, then adds the sucrose of 10g and the 6-benzyl aminoadenine of 0.1 μm ol, obtain culture medium.
Bog moss protonema rapid propagation method the most according to claim 1, it is characterised in that: in step A, magnesium sulfate is MgSO4·7H2O。
Bog moss protonema rapid propagation method the most according to claim 1, it is characterised in that: in step A, iron chloride is FeCl3 ·6 H2O。
Bog moss protonema rapid propagation method the most according to claim 1, it is characterised in that: in step A, culture medium being loaded in 500ml triangular flask, the most bottled about 200ml, autoclaving is stand-by.
Bog moss protonema rapid propagation method the most according to claim 1, it is characterised in that: in step B, bog moss protonema pulverizes 1min at superclean bench, and specific breakage rate is 18000r/min, and 200 mesh cells are sieved through filter；The bog moss protonema crushed is moved in sterilized water, makes suspension.
Bog moss protonema rapid propagation method the most according to claim 1, it is characterised in that: in step C, every 10ml culture medium adds 1ml suspension.
Bog moss protonema rapid propagation method the most according to claim 1, it is characterised in that: in step C, putting into shaking table, rotating speed 140r/min, illumination is 3000lux, and the photoperiod is day night 10 h/ 14 h.
Bog moss protonema rapid propagation method the most according to claim 1, it is characterised in that: in step D, putting into shaking table, rotating speed 140r/min, illumination is 3000lux, and the photoperiod is day night 10 h/ 14 h.
Bog moss protonema rapid propagation method the most according to claim 1, it is characterised in that: in step D, every 10ml culture medium adds 1ml suspension.
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| CN109169021A (en) * | 2018-10-18 | 2019-01-11 | 贵州工程应用技术学院 | A method of it being quickly laid with plantation bog moss on the urban river riparian wetland of plateau |
| CN109588315A (en) * | 2018-12-31 | 2019-04-09 | 西南民族大学 | A kind of sterilization method of bryophyte explant |
| CN113265369A (en) * | 2021-05-25 | 2021-08-17 | 内蒙古大学 | Liquid culture method for rapidly improving biomass of sphagnum brevipedunculatum protonema |
| CN116636463A (en) * | 2023-07-10 | 2023-08-25 | 广西壮族自治区中国科学院广西植物研究所 | Cultivation method of sphagnum moss in warm land in high-altitude area |
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| CN116636463A (en) * | 2023-07-10 | 2023-08-25 | 广西壮族自治区中国科学院广西植物研究所 | Cultivation method of sphagnum moss in warm land in high-altitude area |
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