CN103238459A - Breeding method of multi-spore lingzhi mushroom - Google Patents
Breeding method of multi-spore lingzhi mushroom Download PDFInfo
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Abstract
The invention relates to a breeding method of a multi-spore lingzhi mushroom and belongs to the technical field of genetic breeding of edible mushrooms. The method mainly relates to conventional breeding (lingzhi tissue isolation) with spore mutation breeding in modern biotechnology. The method includes the steps of selection of parents, conventional breeding, induced mutation, variety screening and artificial cultivation. Compared with the prior art, the method has the advantages that sporulation quantity of the variety of multi-spore lingzhi mushroom bred by the breeding method can be increased to 8-16%, more than double of that of general variety of lingzhi mushroom (having the sporulation quantity of 4%), the bred multi-spore lingzhi mushroom is stable in inherited traits and main production traits (sporulation quantity), and accordingly lingzhi mushroom planting is converted from quantity type to quality type, commodity traits of the variety are improved and breeding time of the multi-spore lingzhi mushroom is also shortened.
Description
Technical field
The present invention relates to the selection of a kind of many spores glossy ganoderma, belong to edible mushroom hereditary and selection technical field.
Background technology
Glossy ganoderma (Ganoderma lucidum) is the treasure in motherland's pools of traditional Chinese medicine, have the reputation of " celestial grass ".Pharmacology and clinical research at all times all proves, the effect that glossy ganoderma truly has and prevents and cures diseases, promotes longevity.The Compendium of Material Medica of the Shennong's Herbal of the Eastern Han Dynasty, the famous physician's Li Shizhen (1518-1593 A.D.) of the Ming Dynasty has detailed and sure record to the effect of glossy ganoderma.Modern pharmacology and clinical practice have further confirmed the pharmacological action of glossy ganoderma, and confirm that GL-B is that glossy ganoderma is strengthened the body resistance to consolidate the constitution, strengthening by means of tonics, the main component of promoting longevity.Glossy ganoderma is formally recorded by NF as medicine now, and American Pharmacopeia had formally been included glossy ganoderma in 2005, and it is again the new resource food of state approval simultaneously, has no side effect, can medicine-food two-purpose.
Scientific research shows, the pharmaceutical component of glossy ganoderma is very abundant, wherein effective ingredient can be divided into ten big classes, comprise GL-B, glossy ganoderma polypeptide, triterpenes, 16 seed amino acids (wherein contain seven kinds of human bodies must amino acid), protein, steroid class, mannitol, coumarin glycosides, alkaloid, organic acid (mainly containing fumaric acid), and micro-Ge, P, Fe, Ca, Mn, Zn etc.Glossy ganoderma has dual regulation to human body, and the kind of curing the disease relates to each systems such as cardiovascular and cerebrovascular, digestion, nerve, internal secretion, breathing, motion, and is especially very remarkable to the preventive and therapeutic effect of tumour, liver pathological changes, insomnia and aging.
The range of application of glossy ganoderma is very extensive.See with regard to traditional Chinese medical science angle is dialectical, go into the five internal organs help whole body QI of five ZANG-organs for glossy ganoderma, so the no matter heart, lung, the dirty weakness of liver,spleen,kidney all can be obeyed it.Glossy ganoderma is cured the disease to plant and relates to each systems such as breathing, circulation, digestion, nerve, internal secretion and motion; Contain inside and outside, woman, youngster, each section's disease of face.Strengthen the body resistance to consolidate the constitution glossy ganoderma with regard to its basic reason, strengthen immunologic function, improve the huge effect of Abwehrkraft des Koepers.It is different from general medicine and only plays therapeutic action at certain disease; also being different from general nutritional health food only replenishes the deficiency of nutrition in a certain respect and strengthens; but bidirectional modulation function of human body balance on the whole; the activity of transfer internal body; regulate human body metabolism function; improve the autoimmunity ability, impel whole internal organ or organ function normalization.
Reishi sporule is that glossy ganoderma is in the growth and maturity phase, the extremely small avette reproductive cell that from glossy ganoderma bacterium hole, ejects, spore is filbert to yellowish-brown, and size has only 5.2～6.9 μ m * 8.5～11.2 μ m, be living body biological, be more suitable for the human body stomach and directly absorb.It has condensed the elite of glossy ganoderma, has whole genetic material and the effect of glossy ganoderma.Its contained GL-B, glossy ganoderma polypeptide, triterpenes, amino acid, protein isoreactivity amount of substance according to surveying and determination are 70 times of ganoderma lucidum fruitbody.Yet traditional lucidum variety sporulation quantity lower (sporulation quantity generally only is 4%) is difficult to satisfy people's demand growing to Reishi sporule; And glossy ganoderma goes out the time long (needing some months consuming time) of sesame test, makes many spores glossy ganoderma breeding work make slow progress.
Summary of the invention
At present for the production of lucidum strain divides two classes: a class mainly is the bacterial classification of ganoderma lucidum fruitbody of gathering, and another kind of mainly is the bacterial classification of gathering lucidum spore powder.The objective of the invention is to provides a kind of new lucidum variety hereditary and selection method at the bacterial classification that is mainly used to gather lucidum spore powder, the lucidum variety that selects can make its sporulation quantity reach 8～16%, be general more than 1 times of lucidum variety (sporulation quantity 4%), and the heredity of the many spores that select lucidum variety and the main production traits (sporulation quantity) are stable, thereby realize that glossy ganoderma population plantation is by the transformation of scalar type to mass type, improve the commodity property of glossy ganoderma, shortened many spores glossy ganoderma breeding time simultaneously.
Realize above-mentioned purpose the technical solution used in the present invention
The present invention adopts edible mushroom hereditary and selection technology, specifically relates generally to conventional seed selection (glossy ganoderma tissue isolation), is aided with the spore mutation breeding in the modern biotechnology, carries out the technical research of many spores glossy ganoderma hereditary and selection, mainly comprises following 5 steps:
1) parent selects
Gather lucidum variety best in quality, that sporulation quantity is higher as the cultivation parent of many spores glossy ganoderma from all parts of the country;
2) conventional seed selection
Step 1) is cultivated the parent separate cultivation acquisition mycelium by the glossy ganoderma tissue isolation, through experiment in cultivation, screening obtains the higher lucidum variety of sporulation quantity, and (the glossy ganoderma tissue isolation namely adopts the young flower bud of glossy ganoderma or cap children tissue to separate, and obtains the method for pure mycelia.They have stronger regeneration capacity and keep the ability of parent's kind property, belong to the vegetative propagation category.Tissue separate draw materials extensive, easy and simple to handle, be easy to success, be the easy separating method of glossy ganoderma pure culture);
3) mutagenic treatment
With step 2) the lucidum variety spore that obtains makes spore suspension, handle by ultraviolet mutagenesis: mutagenic treatment should be carried out in camera bellows, dress 15w uviol lamp is 1 in the case, hang on the 30cm eminence, the 20min that should turn on light earlier during mutagenic treatment makes Wavelength stabilizedly, then spore suspension is poured in the sterile petri dish that diameter is 6cm, open the ware lid, irradiation time 1～3min; Dilute postradiation spore suspension more than 1000 times with sterile water, get again on the culture dish that dilution 0.2ml coats the potato agar medium, cultivate 5～10d down at 25 ℃, just can obtain dozens of list bacterium colony at each flat board, select pure, healthy and strong single bacterium colony to move in the slant tube, a bacterium colony only connects an inclined-plane, after treating that the inclined-plane mycelia grows well, insert respectively again in 2～4 bottles of original seeds, carry out experiment in cultivation then, compare the biological property of each bacterium colony repeatedly, leave and take according to qualifications;
4) screening varieties
By step 2) and 3), seed selection-mutagenesis-seed selection again-mutagenesis again filters out the bacterial strain (sporulation quantity height) that meets main production traits requirement, thereby reduces the conventional breeding time, realizes many spores lucidum variety inheritting and selecting work rapidly and efficiently;
5) artificial cultivation
The improved seeds that will obtain by step 4) carry out artificial cultivation, realize the fast growing (sporulation quantity reaches 8～16%) of many spores glossy ganoderma; Kind heredity and the main production traits (sporulation quantity) are stable, have shortened many spores glossy ganoderma breeding time simultaneously.
Beneficial effect of the present invention
The present invention adopts edible mushroom hereditary and selection technology, specifically relates generally to conventional seed selection (glossy ganoderma tissue isolation), is aided with the spore mutation breeding in the modern biotechnology, carries out the technical research of many spores glossy ganoderma hereditary and selection.Filter out improved seeds, realize the fast growing (sporulation quantity reaches 8～16%) of many spores glossy ganoderma; Kind heredity and the main production traits (sporulation quantity) are stable.Many spores glossy ganoderma selection provided by the invention can scale, industrialization; for the genetic breeding of glossy ganoderma has been opened up a new road; thereby realize that glossy ganoderma plantation scalar type to the transformation of mass type, improves the commodity property of kind, shortened many spores glossy ganoderma breeding time simultaneously.
Embodiment
Embodiment 1
The glossy ganoderma harvesting standard: disappear when fruit body cap edge is yellow, come to the ripening period, this phase cap is no longer expanded, but can continue to increase weight, thicken, and suitably control is ventilated, and the effect of the sporophore growth of promotion is arranged.Disappear when fruit body cap edge is yellow, gradually during overstrike, can gather.
The glossy ganoderma collecting method is as follows:
1) collecting time: August～September/every year;
2) collecting method: when glossy ganoderma is gathered, cut off from the following 3cm of sesame lid position with fruittree shear, stay stem in order to regenerate the second damp glossy ganoderma;
3) points for attention: when glossy ganoderma is gathered, can not hold cap, in order to avoid the cap wound causes infection: the rainy day must not gather, and gathers every 2 days at least after raining heavily.To gently adopt in the glossy ganoderma recovery process and put down gently, prevent from damaging, weigh wounded and affix sandy soil and impurity.
Embodiment 2
The glossy ganoderma tissue separates to be cultivated: select the Guizhou clean higher lucidum variety of mountain sporulation quantity of ancient India, adopt the young flower bud of glossy ganoderma or cap children tissue to separate cultivation, obtain pure mycelia; They have stronger regeneration capacity and keep the ability of parent's kind property, and concrete operations are as follows:
1) young flower bud separates: cut the young flower bud that glossy ganoderma stem tip expands, under aseptic condition, with 75% cotton ball soaked in alcohol wiping, again with the blade after the calcination from shallow 1 cutter of cutting of shank, break into two with one's hands behind the two halves to choose with sharp vaccinating lancet and cut bacterial context 1 fritter in the young flower bud, move and be connected to inclined-plane central authorities, 28 ℃ of following shadings are cultivated, and mycelia is covered with the inclined-plane and is female the kind;
2) cap separates: choose normal, the healthy and strong and immature fruit body of growth, cut stem, with 75% cotton ball soaked in alcohol wiping cap positive and negative, cot is cut brokenly at the yellow growth circle place at the cap edge then, break down 1 of young tender cap off with the fingers and thumb, with the scalpel after the calcination section bacterial context is cut into the tissue grain of grain of rice size in length and breadth, put in the sterile petri dish, will organize grain to move with transfer needle at last and be connected on the inclined-plane, 28 ℃ of following shadings are cultivated, and whole operation needs to carry out under strict aseptic condition.
Embodiment 3
Mutation breeding: select the Guizhou clean higher lucidum variety of mountain sporulation quantity of ancient India to carry out mutation breeding, mutagen are ultraviolet ray, and mutation breeding comprises following four steps:
1) collection of the aseptic spore of fresh glossy ganoderma: many covers culture dish, glass funnel and stainless steel hook with sterilization treatment is crossed place on the superclean bench; The ganoderma lucidum fruitbody cap of selecting to launch spore is with 75% cotton ball soaked in alcohol wiping, under the sterile working state, it is hung in the glass funnel, after ganoderma lucidum fruitbody launches naturally through 12～20h, can collect the aseptic spore of fresh glossy ganoderma that obtains some in the culture dish;
2) preparation of spore suspension: the aseptic spore of the glossy ganoderma that collection is obtained moves in stroke-physiological saline solution or the phosphate buffer, shake up the back spore suspension;
3) mutagenic treatment: use UV treatment, mutagenic treatment should be carried out in camera bellows, dress 15w uviol lamp is 1 in the case, hang on the 30cm eminence, 20min should turn on light earlier during mutagenic treatment, make Wavelength stabilized, spore suspension is poured in the sterile petri dish that diameter is 6cm then, opened the ware lid, irradiation time 1～3min, dilute postradiation spore suspension more than 1000 times with sterile water, get then on the culture dish that dilution 0.2ml coats the potato agar medium, cultivate 5～10d down at 25 ℃, at this moment just can obtain dozens of list bacterium colony at each flat board, select pure, healthy and strong single bacterium colony moves in the slant tube, for further screening;
4) choose the bacterium screening: the spore after the mutagenesis is in time chosen bacterium after cultivation grows bacterium colony, publish and educate in the sound single bacterium colony immigration slant tube, a bacterium colony only connects an inclined-plane, after treating that the inclined-plane mycelia grows well, insert respectively again in 2～4 bottles of original seeds, carry out experiment in cultivation then, compare the biological property of each bacterium colony repeatedly, leave and take according to qualifications.
Embodiment 4
Artificial cultivation: with the above-mentioned improved seeds that obtain by " seed selection-mutagenesis-seed selection again-mutagenesis again ", carry out artificial cultivation, realize the fast growing of many spores glossy ganoderma.Should note following item in the artificial cultivation process:
1) do the sanitation and hygiene of planting environment well: will remove bacterium slag, rubbish in the sesame room, thoroughly clean cultivation and use frame, assorted bacterium concealment place is eliminated in the row space of going forward side by side sterilization, reduces communication media, does assanation well and can play a multiplier effect to preventing pollution;
2) strictness is selected materials: the composts or fertilisers of cultivating requirement is fresh, nothing is gone mouldy, with before to tan by the sun a couple of days, the composts or fertilisers of cultivating prescription requires rationally, major ingredient and auxiliary material will fully be mixed thoroughly, water content control is about 60%～70%, and loading amount is suitable, degree of tightness is appropriate, carries out high pressure or normal-pressure sterilization after installing immediately, in case the acidifying of medium, sterilization requires thoroughly;
3) establish strict aseptic concept in the inoculation: owing to floating in the air assorted bacterium spore arranged everywhere, can not be off one's guard because naked eyes cannot see during operation, operating personnel's both hands, clothing and used inoculating tool, material palpus strict sterilization, as select for use the operating personnel of transfer room inoculation to put on a hat, in case fall to having assorted bacterium spore on the hair.Inoculation operation will try one's best fast, skilled, prevent from bringing in the seeded process assorted bacterium, assorted bacterium spore, sack damaged in the sterilization process is sealed with adhesive plaster, and at sealing part 75% alcohol disinfecting;
4) suitably strengthen inoculum concentration: can make Ganoderma lucidum mycelium capture the domain rapidly with absolute predominance, reduce the pollution of assorted bacterium, play with the antibacterial effect of sesame;
5) guarantee to have suitable microclimate in the culturing room, holding cultural hypha closes: control about 25 ℃ temperature, 60%～70% humidity, note ventilation, be strictly on guard against hot and humid, create the optimum environmental condition of Ganoderma lucidum mycelium growth, promote Ganoderma lucidum mycelium to grow fast, capture whole charge level rapidly;
6) conscientiously do a good job of it out the cultivation management work in sesame stage: cultivation of glossy ganoderma generally is selected in and carries out spring, when going out sesame just in time is 6, the high temperature season in July, and therefore the sporophore growth stage is the important period of control living contaminants owing to need higher humidity.After the glossy ganoderma original hase grows, in time take out tampon or open bag, in order to avoid original hase damages and bacteria infection.Carry out heat and moisture preserving work, add forced ventilation simultaneously and give certain illumination, impel original hase to grow up to fruit body healthily, in time pluck after the fruit body maturation;
7) strengthen early prevention and treatment: make regular check on growing state, in case find to pollute, should adopt an effective measure, the growth of the assorted bacterium of control, spread.Generally can adopt limewash to clean the affected part, or close down with lime white; As the fruit body bacteria infection, should in time extract, in case spread.Can select also in cultivation that some are efficient for use, the bactericide of low toxicity, low-residual carries out spice or treatment, but must be prudent when selecting bactericide for use, in order to avoid kill mycelia, or cause residue of pesticide.
Many spores lucidum variety that this method selects can make its sesame sporulation quantity reach 8～16%, be general more than 1 times of lucidum variety (sporulation quantity 4%), and the many spores that select glossy ganoderma is stable in heredity and the main production traits (sporulation quantity), thereby realize that the glossy ganoderma artificial planting is changed to mass type by scalar type, improve the commodity property of kind, shortened many spores glossy ganoderma breeding time simultaneously.
Claims (2)
1. the selection of spore glossy ganoderma more than a kind, it is characterized in that: it is the glossy ganoderma tissue isolation that this method adopts conventional seed selection, is aided with the spore mutation breeding in the modern biotechnology, carries out the technical research of many spores glossy ganoderma hereditary and selection, mainly comprises following 5 steps:
1) parent selects
Gather lucidum variety best in quality, that sporulation quantity is higher as the cultivation parent of many spores glossy ganoderma from all parts of the country;
2) conventional seed selection
Step 1) is cultivated the parent separate the pure mycelium of cultivation acquisition by the glossy ganoderma tissue isolation, through experiment in cultivation, screening obtains the higher lucidum variety of sporulation quantity---the method for glossy ganoderma tissue branch
Adopt the children of glossy ganoderma
Or bacterium
Children's tissue
Separate and cultivate, obtain the method for pure mycelia:
3) mutagenic treatment
With step 2) the lucidum variety spore that obtains makes spore suspension, handle by ultraviolet mutagenesis, with sterile water the postradiation spore suspension of ultraviolet ray is diluted, getting dilution again coats on the culture dish of potato agar medium, cultivate 5～10d down at 25 ℃, obtain single bacterium colony at flat board, select pure, healthy and strong single bacterium colony to move in the slant tube, a bacterium colony only connects an inclined-plane, after treating that the inclined-plane mycelia grows well, insert respectively again in 2～4 bottles of original seeds, carry out experiment in cultivation then, compare the biological property of each bacterium colony repeatedly, leave and take according to qualifications;
4) screening varieties
By step 2) and 3), seed selection-mutagenesis-seed selection again-mutagenesis again filters out the high bacterial strain of sporulation quantity that meets main production traits requirement, thereby reduces the conventional breeding time, realizes many spores lucidum variety inheritting and selecting work rapidly and efficiently;
5) artificial cultivation
The improved seeds that will obtain by step 4) carry out artificial cultivation, realize the fast growing of many spores glossy ganoderma, and its sporulation quantity reaches 8～16%; Kind heredity and main sporulation quantity proterties are stable, have shortened many spores glossy ganoderma breeding time simultaneously.
2. according to the selection of the described a kind of many spores glossy ganoderma of claim 1, it is characterized in that:
Step 2) concrete operations of glossy ganoderma tissue isolation comprise: A. children flower bud separates: cut the young flower bud that glossy ganoderma stem tip expands, under aseptic condition, with 75% cotton ball soaked in alcohol wiping, again with the blade after the calcination from shallow 1 cutter of cutting of shank, break into two with one's hands behind the two halves to choose with sharp vaccinating lancet and cut bacterial context 1 fritter in the young flower bud, move and be connected to inclined-plane central authorities, 28 ℃ of following shadings are cultivated, and mycelia is covered with the inclined-plane and is female the kind; B. cap separates: choose normal, the healthy and strong and immature fruit body of growth, cut stem, with 75% cotton ball soaked in alcohol wiping cap positive and negative, cot is cut brokenly at the yellow growth circle place at the cap edge then, breaks down 1 of young tender cap off with the fingers and thumb, the section bacterial context is cut in length and breadth the tissue grain of grain of rice size with the scalpel after the calcination, to organize grain to move with transfer needle again and be connected on the inclined-plane, 28 ℃ of following shadings are cultivated, and whole operation needs to carry out under strict aseptic condition.
The concrete operations of step 3) mutagenic treatment comprise: the collection of the aseptic spore of the fresh glossy ganoderma of A.: many covers culture dish, glass funnel and stainless steel hook with sterilization treatment is crossed place on the superclean bench; The ganoderma lucidum fruitbody cap of selecting to launch spore is with 75% cotton ball soaked in alcohol wiping, under the sterile working state, it is hung in the glass funnel, after ganoderma lucidum fruitbody launches naturally through 12～20h, can collect the aseptic spore of fresh glossy ganoderma that obtains some in the culture dish; B. the preparation of spore suspension: the aseptic spore of fresh glossy ganoderma is moved in stroke-physiological saline solution or the phosphate buffer, shake up the back spore suspension; C. mutagenic treatment: use UV treatment, mutagenic treatment should be carried out in camera bellows, dress 15w uviol lamp is 1 in the case, hang on the 30cm eminence, 20min should turn on light earlier during mutagenic treatment, make Wavelength stabilized, then spore suspension is poured in the sterile petri dish that diameter is 6cm, open the ware lid, irradiation time 1～3min dilutes postradiation spore suspension more than 1000 times with sterile water, get then on the culture dish that dilution 0.2ml coats the potato agar medium, cultivate 5～10d down at 25 ℃, select pure, healthy and strong single bacterium colony moves in the slant tube, for further screening; D. choose the bacterium screening: the spore after the mutagenesis is in time chosen bacterium after cultivation grows bacterium colony, publish and educate in the sound single bacterium colony immigration slant tube, a bacterium colony only connects an inclined-plane, after treating that the inclined-plane mycelia grows well, insert respectively again in 2～4 bottles of original seeds, carry out experiment in cultivation then, compare the biological property of each bacterium colony repeatedly, leave and take according to qualifications.
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN104956914A (en) * | 2015-06-18 | 2015-10-07 | 唐伟 | Breeding method for natural ganoderma lucidum |
| CN115024159A (en) * | 2022-06-23 | 2022-09-09 | 海南热带海洋学院 | Method for screening heat-resistant ganoderma lucidum variety |
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| CN102119631A (en) * | 2010-12-09 | 2011-07-13 | 江苏大学 | Grifola frondosa strain for producing polysaccharide with composite raw material of rice bran and wheat bran |
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| CN1631078A (en) * | 2003-12-23 | 2005-06-29 | 马士贵 | Wild-imitating mushroom edible fungus seed and cultivation method thereof |
| CN102119631A (en) * | 2010-12-09 | 2011-07-13 | 江苏大学 | Grifola frondosa strain for producing polysaccharide with composite raw material of rice bran and wheat bran |
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| CN115024159A (en) * | 2022-06-23 | 2022-09-09 | 海南热带海洋学院 | Method for screening heat-resistant ganoderma lucidum variety |
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Application publication date: 20130814 |