CN103650912B - A kind of multi-spore inbred breeding method for Flammulina velutipes - Google Patents
A kind of multi-spore inbred breeding method for Flammulina velutipes Download PDFInfo
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Abstract
The invention belongs to the Breeding of Edible Mushroom technical field, be specifically related to a kind of multi-spore inbred breeding method for Flammulina velutipes.Utilize the breeding seed selection Asparagus novel bacterial of liquid culture Quito spore selfing.The steps include: that (1) is collected single spore and made spore liquid from the fruit body in parent strain Gaoyou 738; (2) shaking bacterium by adding in liquid nutrient medium containing the spore liquid of parent strain, carrying out the selfing of many spores; (3) liquid spawn Shaking culture is carried out mycelium stimulation process; (4) aimed strain Asparagus (Flammulina velutipes) D7-Y is obtained by cultivating and screening.Compared with prior art, the present invention have be applicable to batch production, simply, feature fast, and pollute the effect being easily found to improve breeding efficiency.
Description
Technical field
The invention belongs to the Breeding of Edible Mushroom technical field, be specifically related to a kind of multi-spore inbred breeding method for Flammulina velutipes.
Background technology
China's Flammulina velutipes germplasm resources enriches.Along with the development of technology, be no matter that the material of the production of hybrid seeds, technique or the breeding technique of bacterial classification are obtained for and improve.Now, factory culture Asparagus will become the main flow in epoch, can a kind of high yield of seed selection, high-quality, high resistance, bioactive substance content is high, mouthfeel good and the bacterial classification of applicable factory culture is most important.
In Asparagus factorial praluction, often find, an original excellent kind is after production after a while, and its merit starts to show not obvious, and the such as fruiting phase obviously postpones, and become mushroom rate low, fruiting is irregular, production declining etc.The various growth factor of manual control and the growth factor of Asparagus the most suitable growth environment variant.The change of environment, can accelerate the variation of some proterties of kind, causes some merit slowly to disappear.And present cultivar used is mostly White strain, and it mostly is hybridization or mutational variety, though this strain each side proterties is all more excellent, its gene mostly are heterozygosity, namely show as dominance geneticing character.According to genetic consideration, certain gene changes, and its proterties just likely changes.So in process of production, Needle mushroom strain especially easily morphs and loses original merit.
Frequently go down to posterity to a kind, unfavorable condition of culture and storage conditions be the main external cause causing failing, causing mycelia genetic material to there occurs variation thus, is the internal cause of bacterial classification decline.Edible fungus species extends along with incubation time, aging phenomenon will occur, and produces a series of physiological acoustic signals.Aging bacterial classification easily occurs that mycelium germination rate declines, growth rate slows down, growing way weakens, lighten, fruit-body formation are delayed, small and weak etc.
Asparagus is China's factorial praluction edible mushroom kind with fastest developing speed.The improvement of kind and seed selection are the demands of industry development always.The breeding technique of current Asparagus mainly contains domesticated breeding, mutation breeding, list-single crosses, list-double cross, many spore hybridization, the methods such as the selfing of many spores and transgenic technology breeding, the most widely used various methods being still crossbreeding, only huge with single double cross workload, outlet (the Xu Zhen etc. of many spore hybridization and the selfing of many spores, 2007) efficiency of breeding is substantially increased, conventional many spores selfing refers to the spore Mixed culture of the fruit body that mother plants being launched in gnotobasis, timely picking hybrid strain afterwards, can effectively get rid of conidial interference, but the culture bottle in factorial praluction is not suitable for accessing solid spawn, need that Hybridizing colonies is made liquid spawn and access culture bottle again, this just makes breeding step increase, the breeding cycle length of side, reduce efficiency on the contrary, so many spores selfing technology still has very large improvement and bring new ideas space.
Summary of the invention
The object of the invention is to the defect overcoming prior art existence, technical problem to be solved is to provide a kind of Asparagus many spores cross breeding method, and the method can be applicable to the seed selection of the excellent new strains of industrialized cultivation for needle mushroom.
The present invention is realized by following technical step:
A kind of Asparagus many spores cross breeding method, its step is as follows:
(1) full grown fruit body cap part is got from Asparagus Gaoyou 738 bacterial strain, cut away stem, cotton ball soaked in alcohol with 75% cleans cap surface, lamella catches on sterilized cleek down and hangs in the triangular flask of sterilizing, spend the night with aseptic tampon sealing, collect spore print, make spore liquid with sterile water;
(2) spore liquid of upper step gained is joined in liquid nutrient medium carry out shaking bacterium, carry out the selfing of liquid many spores, shaking bacterium condition is: rotating speed 140r/min, cultivation temperature is 25 DEG C, when the limpid mycelium pellet of culture fluid enriches time, shake bacterium terminate, stir evenly mycelium pellet with magnetic stirring apparatus and obtain Asparagus mixing mycelia liquid;
(3) Asparagus of upper step gained mixing mycelia liquid is being inoculated on the composts or fertilisers of cultivating of culture bottle in toilet, with ventilating cover sealing, and light culture 25-30d in the culturing room being placed in 14-15 DEG C;
(4) cover with after culture bottle until mycelia, strike off bottleneck top layer mycelia and carry out mycelium stimulation, to stimulate fruiting;
(5) culture bottle after mycelium stimulation is moved into fertility room, initial temperature 18 DEG C, successively decreases by number of days, finally remain on 4 DEG C to cultivate, screening to be less than that 25d, mushroom shape are neat, attractive in appearance breeding time, stem base portion without the bacterial strain of fine hair, the overall consolidation of bacterium colony, carry out tissue separation, obtain candidate strain; Multiple cropping is carried out to obtained candidate strain, by being less than breeding time, 25d, mushroom shape are neat, attractive in appearance, stem base portion is greater than 350g without fine hair, the overall consolidation of bacterium colony and single bottle of output and carries out multiple sieve once, obtain Strains of Flammulina velutipes D7-Y, Flammulina velutipes D7-Y, China is sent by this bacterial strain on November 24th, 2013. Wuhan. Wuhan University's China typical culture collection center preservation, preserving number is CCTCC NO:M2013599.
Wherein:
Liquid culture based formulas is as follows:
Soybean meal 3.3g/L, sucrose 20g/L, epsom salt 0.66g/L, hydrogen sulfate dipotassium 0.66g/L, adds water and is settled to 1L.pH=6.2-6.5。
The culture material formula of culture bottle is made up of the raw material of following weight percentage:
The fruit body feature of Strains of Flammulina velutipes D7-Y:
The shallow milk yellow of stem, cap is yellow, fruiting temperature 4 ~ 17 DEG C, mushroom handle is long stiffening, and mushroom handle top is light yellow, and root is yellow, without brown, mushroom lid is involute, neatly, more not easily parachute-opening, fruiting is close together, and color and luster is glossy, low temperature resistant, output is higher, a tide of only gathering in biological conversion rate 110%(factory), breeding time short (relative factory cultivar shifts to an earlier date 5-10d), be comparatively applicable to factory culture.
Compared with prior art, good effect of the present invention is:
1, the present invention substantially reduces the breeding cycle of Asparagus.
2, pollution is more easily found, and improves breeding efficiency.
3, operating procedure of the present invention is simple, and the needs of liquid spawn are prepared in applicable batch production.
More detailed technical scheme is shown in " embodiment ".
Accompanying drawing explanation
Fig. 1: be general technical route map of the present invention.
Fig. 2: be the Asparagus new strains D7-Y fruit body feature photo that the present invention obtains.
Embodiment
Embodiment 1:
The present embodiment parent strain used for Gaoyou 738 for science edible mushroom research institute of Gaoyou City of Jiangsu Province provides (see download address: Chinese agriculture net: http://www.zgny.com.cn/ProHtml/5/1/5/15638.html).
Full grown fruit body cap part is got from Asparagus Gaoyou 738 bacterial strain, cut away stem, cotton ball soaked in alcohol with 75% cleans cap surface, lamella catches on sterilized cleek down and hangs in the triangular flask of sterilizing, spend the night with aseptic tampon sealing, collect spore print, make spore liquid with sterile water;
Asparagus Gaoyou 738 spore liquid of upper step is directly joined (additional proportion: add 0.5ml spore liquid in 600ml culture fluid in triangle shaking flask, 300ml culture fluid adds 0.2-0.3ml spore liquid), carry out the selfing of many spores (at rotating speed 140r/min shaking table, temperature is carry out Shaking culture in the culturing room of 25 DEG C, obtain many spores selfing culture fluid in Gaoyou 738), terminating for shaking bacterium when the limpid mycelium pellet of culture fluid enriches time, stirring evenly mycelium pellet with magnetic stirring apparatus and obtaining Asparagus mixing mycelia liquid;
Upper step gained Asparagus mixing mycelia liquid is directly inoculated into (many spores selfing culture fluid in every bottle graft kind 50ml Asparagus Gaoyou 738) in culture bottle, with ventilating cover sealing, and light culture 25-30d in the culturing room being placed in 14-15 DEG C;
Cover with after culture bottle until mycelia, strike off bottleneck top layer mycelia and carry out mycelium stimulation, to stimulate fruiting;
Culture bottle after mycelium stimulation is moved into fertility room (initial temperature 18 DEG C, successively decrease by number of days, finally remain on 4 DEG C) cultivate, screening to be less than that 25d, mushroom shape are neat, attractive in appearance breeding time, stem base portion is without the bacterial strain of fine hair, the overall consolidation of bacterium colony, carry out tissue to be separated, obtain candidate strain, multiple cropping is carried out to obtained candidate strain, by being less than breeding time, 25d, mushroom shape are neat, attractive in appearance, stem base portion is greater than 350g without fine hair, the overall consolidation of bacterium colony and single bottle of output and carries out multiple sieve once, final acquisition one strain strain excellent, this Strain Designation is Asparagus D7-Y by applicant; Flammulina velutipes D7-Y; On November 24th, 2013, this bacterial strain is delivered China. Wuhan. Wuhan University's China typical culture collection center (CCTCC) preservation, its preserving number is CCTCCNO:M2013599.
Aforesaid liquid culture medium prescription is as follows:
Soybean meal 3.3g/L, sucrose 20g/L, epsom salt 0.66g/L, hydrogen sulfate dipotassium 0.66g/L, adds water and is settled to 1L.pH=6.2-6.5。
The culture material formula of culture bottle is made up of the raw material of following weight percentage:
Table 1 different breeding method batch production seed selection Asparagus correlation coefficient compares
As can be seen from Table 1, the present invention can shorten breeding cycle greatly, can pollute and take measures by Timeliness coverage, improve breeding efficiency.
Leading reference:
1. Xu Zhen etc., the Needle mushroom strain of many spores selfed breeding factory culture, fungus journal 2007(4).
Claims (3)
1. Asparagus many spores cross breeding method, is characterized in that the following step:
(1) full grown fruit body cap part is got from Asparagus Gaoyou 738 bacterial strain, cut away stem, cotton ball soaked in alcohol with 75% cleans cap surface, lamella catches on sterilized cleek down and hangs in the triangular flask of sterilizing, spend the night with aseptic tampon sealing, collect spore print, make spore liquid with sterile water;
(2) spore liquid of upper step gained joined in liquid nutrient medium and carry out shaking bacterium, carry out the selfing of liquid many spores, shaking bacterium condition is: rotating speed 140r/min, and cultivation temperature is 25 DEG C, obtains Asparagus mixing mycelia liquid;
(3) Asparagus of upper step gained mixing mycelia liquid is inoculated on the composts or fertilisers of cultivating of culture bottle in toilet, with ventilating cover sealing, and light culture 25-30d in the culturing room being placed in 14-15 DEG C;
(4) cover with after culture bottle until mycelia, strike off bottleneck top layer mycelia and carry out mycelium stimulation, to stimulate fruiting;
(5) culture bottle after mycelium stimulation is moved into fertility room, initial temperature 18 DEG C, successively decreases by number of days, finally remain on 4 DEG C to cultivate, screening to be less than that 25d, mushroom shape are neat, attractive in appearance breeding time, stem base portion without the bacterial strain of fine hair, the overall consolidation of bacterium colony, carry out tissue separation, obtain candidate strain; Multiple cropping is carried out to obtained candidate strain, by being less than breeding time, 25d, mushroom shape are neat, attractive in appearance, stem base portion is greater than 350g without fine hair, the overall consolidation of bacterium colony and single bottle of output and carries out multiple sieve once, obtain the Asparagus D7-Y bacterial classification that preserving number is CCTCC NO:M 2013599;
Wherein:
Liquid culture based formulas:
Soybean meal 3.3g/L, sucrose 20g/L, epsom salt 0.66g/L, hydrogen sulfate dipotassium 0.66g/L, adds water and is settled to 1L, pH=6.2-6.5;
The culture material formula of culture bottle is made up of the raw material of following weight percentage:
By dry basis percentage by weight:
2. one to select good strains in the field for seed the Asparagus D7-Y bacterial classification of educating, be deposited in China typical culture collection center, preserving number is CCTCC NO:M 2013599.
3. the application of Asparagus D7-Y bacterial classification according to claim 2 in Asparagus high-yield culturing.
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| CN104221709B (en) * | 2014-08-30 | 2016-08-17 | 永州市祥瑞生物科技有限公司 | A kind of production method of Needle mushroom strain |
| CN105349436B (en) * | 2015-12-05 | 2019-05-07 | 中农伟业(北京)生态科技发展有限公司 | Wild matsutake tames parent species preparation method |
| CN107142257B (en) * | 2017-05-19 | 2020-10-09 | 山西农业大学 | Breeding method of light yellow needle mushrooms |
| CN112831456A (en) * | 2019-11-25 | 2021-05-25 | 湖南金芙农业科技有限公司 | Tricholoma matsutake sexual spore and separation method thereof |
| CN111518708A (en) * | 2020-01-03 | 2020-08-11 | 福建农林大学 | Yellow needle mushroom strain nong jin 49 not easy to open and molecular marker identification method thereof |
| CN115136847A (en) * | 2022-07-07 | 2022-10-04 | 陕西春森菌业有限公司 | Edible fungus spore seed production technology with fungus curtain |
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| CN102396343A (en) * | 2011-04-22 | 2012-04-04 | 上海市农业科学院 | Method for fruiting and breeding needle mushrooms on flat plate through multi-spore self-breeding |
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