CN104285815A - Tissue culture and rapid propagation method of E. urophylla*E. Grandis DH32-13 - Google Patents
Tissue culture and rapid propagation method of E. urophylla*E. Grandis DH32-13 Download PDFInfo
- Publication number
- CN104285815A CN104285815A CN201410590691.5A CN201410590691A CN104285815A CN 104285815 A CN104285815 A CN 104285815A CN 201410590691 A CN201410590691 A CN 201410590691A CN 104285815 A CN104285815 A CN 104285815A
- Authority
- CN
- China
- Prior art keywords
- culture
- days
- tissue culture
- seedling
- explant
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
Classifications
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A40/00—Adaptation technologies in agriculture, forestry, livestock or agroalimentary production
- Y02A40/10—Adaptation technologies in agriculture, forestry, livestock or agroalimentary production in agriculture
- Y02A40/22—Improving land use; Improving water use or availability; Controlling erosion
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02P—CLIMATE CHANGE MITIGATION TECHNOLOGIES IN THE PRODUCTION OR PROCESSING OF GOODS
- Y02P60/00—Technologies relating to agriculture, livestock or agroalimentary industries
- Y02P60/40—Afforestation or reforestation
Landscapes
- Breeding Of Plants And Reproduction By Means Of Culturing (AREA)
Abstract
The invention discloses a tissue culture and rapid propagation method of E. urophylla*E. Grandis DH32-13. The method is implemented through the steps of explant establishment, explant disinfection, primary culture, subculture, rooting culture, seedling-hardening, transplantation and the like. According to the invention, special culture medium components are selected for the primary culture, subculture and rooting culture, the E. urophylla*E. Grandis is fast in growth and high in survival rate after being transplanted by virtue of a reasonable seedling-hardening measure, and the purposes of high propagation coefficient and excellent propagation effect are achieved; besides, the method is low in production cost, and the genetic characteristics of obtained tissue culture seedlings are consistent, therefore, the method is an effective way for tissue culture and rapid propagation of eucalyptuses, and has wide application range.
Description
Technical field
The present invention relates to Eucalyptus Planting technical field, particularly a kind of tissue culture and rapid propagation method of tail alpine ash DH32-13 kind.
Background technology
Eucalyptus (Eucalyptus) is the general designation that Myrtaceae (Myrtaceae) eucalyptus belongs to (Eucalyptus) plant, originate in Australia, through the introduction and acclimatization in country variant for many years and area, develop into more than 600 kind, wherein Important Economic commerical tree species about more than 100 are planted, and are had kind more than 40 by what extensively cultivate.In recent years, eucalyptus plantation is all being greatly developed in China various places, and in Guangdong and Guangxi Provinces, Hainan, Sichuan, Yunnan etc. economizes and take on a certain scale, and creates significantly ecological and economic benefit.
Tail alpine ash DH32-13 kind (
e. urophylla × E. Grandis), be forest farm, east gate, Guangxi, with Indonesia's provenance east gate Eucalyptus urophylla select tree U16(
eucalyptus urophylla) as maternal, with Australian provenance east gate alpine ash select tree G46(
eucalyptus grandis) as the heterosis, hybrid vigor kind of male parent, by the excellent eucalypt species that vegetative propagation obtains, this kind has fast growing, rotation short (industrial fiber material); Form is logical directly, branch is little, forest form is neat; Material is good, and texture is straight, barren-resistant, be applicable to the advantage of large area region plantation, all can plant in each department, Guangxi, and extend trial for many years prove, 5 years raw tail alpine ash DH32-13 kind mean stand height 24.52m, mean DBH increment 17.7cm, average per hectare accumulation reaches 45m
3/ hm
2above, economy and ecological benefits are very remarkable, and by excellent kind amount reproduction at short notice, acquired character performance is consistent, genetic stability, the offspring that trait segregation does not occur is significant, eucalyptus is the woody plant of perennial cross pollination, the phenomenon that between kind, natural hybrization hybridizes is frequent, offspring is seriously separated, therefore adopt sexual propagation means to be difficult to keep select tree characteristic, and asexual reproduction method such as cuttage, press strip, grafting etc. are unfavorable for amount reproduction, it is comparatively effective method that tissue culture technique cultivates Fast-propagation eucalyptus.
Tissue cultures (Tissue Culture) is aseptically, and be separated also the technology of culture of ex vivo plant tissue in the medium, it has following significance: (1) promotes the Fast-propagation of excellent genetic stocks; (2) cultivation of detoxification breeding seedling and the amount reproduction of virus-free seedling; (3) Fast-propagation of special breeding material; (4) Fast-propagation of gene engineering plant; (5) Fast-propagation of Plantlet in vitro kind matter.According to statistics, current existing more than 60 plant eucalyptus tissue cultures succeeds, first reported eucalyptus camaldulensis seedling explant obtains callus from the Susses sixties in last century, both at home and abroad successively in eucalyptus citriodora (1969), alpine ash (1974), beautiful flower eucalyptus (1976), Bai An (1975), hybrid eucalyptus (1980), grey eucalyptus, Wang An (1980), Lei Lin mono-eucalyptus (1981), willow gallery F1 hybridizes generation (1981), eucalyptus camaldulensis (1992) has carried out linked groups's culture technique research.
Meanwhile, the research for the tissue cultures aspect of eucalyptus also has many:
1, title: the stereoscopic culture of tail alpine ash and Fast-propagation; Author: Liu Yiqing, Wang Daping, Xiong Yunhai; Gardening journal the 32nd volume; Summary: the tissue culture and rapid propagation method discussing No. T13, tail alpine ash.Gather the semi-lignified spray of No. T13, tail alpine ash in the season of growth, remove blade, be cut into stem-segment with single bud, use tap water 15min, 0.5% washing powder solution soaks 15min, cleaner with tap water.Material is put into respectively the empty cup of aseptic plastic, put 15-20,75% alcohol-pickled 30s for every glass, 0.1% mercuric chloride sterilization 10-12min, finally uses aseptic water washing 3-5 time, is inoculated in inducing culture.The minimal medium of induction axillary bud sprouting and proliferation and subculture is kept to 1/3 for improvement MS(ammonium nitrate, is called for short EU), the minimal medium of taking root is 1/2MS, BA, the IBA of additional variable concentrations, NAA, active carbon (AC), sucrose 3%, agar 5.0g./L, pH6.0.Fiber differentiation utilizes Indoor Natural scattered light (not turning on light), and proliferation and subculture, strengthening seedling and rooting are cultivated, illumination 12h/d, light intensity 1500-2000lu, temperature 26 ± 3 DEG C, ambient humidity 405-70%.Results and analysis: EU+BA0.5+NAA0.2 evoked callus is best, and cultivation effect is best.This research uses traditional sucrose as carbon source, and culture medium cost is relatively high, and the nutrient component of sucrose is single, more comprehensively for cell division provides nutriment, can not affect culture effect to a certain extent.In addition, do not add the composition improving resistance and suppress living contaminants in medium, the survival rate of plantlet in vitro can be affected.
2, title: eucalyptus tissue culture rapid propagating technology; Author: Lin Yingyan; Guangxi Vocational & Technical College's thesis (design), summary: more detailed discuss the formula (MS+BA0.5+IBA0.2) of eucalyptus from the sterilizing of the sampling of Initial culture, material, inoculation, cultivation and medium; Inoculation method in subculture, medium (MS+BA1.0+KT0.5+IBA0.1-0.5) and condition of culture; The formula (1/2MS+ABT1.5+IBA0.1+ active carbon 0.25%) of the medium in culture of rootage, inoculation method and cultivation and condition; Various control measures etc. in hardening in nursery, the preparation of matrix, transplanting and seedling raising process.Equally, do not add the composition improving resistance and suppress living contaminants in the medium of this research, the survival rate of plantlet in vitro can be affected.
3, Authorization Notice No. is the patent of CN103155872B: a kind of tail alpine ash embryoid induction seedling-cultivating method, and it for explant with tail alpine ash aseptic seedling stem sections, is sprouted through embryonic callus induction, embryoid induction and embryoid and can obtain complete regeneration plant; Adopt method of the present invention can obtain through embryoid way regeneration plant, result is stablized, and repeatability is good, can apply and produce upper large-scale commercial nursery, for the breeding of tail alpine ash is laid a good foundation.Above-mentioned patent adopts tail alpine ash aseptic seedling stem sections to be that explant carrys out induced embryonic callus, regeneration plant is obtained by somatic embryo generation Regeneration Ways, operation is comparatively complicated, enforcement difficulty is large, use tail alpine ash aseptic seedling stem sections to be explant simultaneously, maternal good characteristic needs to be investigated, and simple this technology commercialization nursery of employing can not effective hereditary parent's good characteristic.
Visible, although more for the research of eucalyptus tissue culture technique at present, but it is numerous that the purposive merit for select tree carries out expansion, the tissue culture and rapid propagation method simultaneously can effectively applied also has a lot of need of work to grope, because DH32-13 is perennial tail alpine ash kind, market plantlet in vitro is less, resource is well sold and in short supply, and for this kind tissue cultures work and not yet in effectly to carry out, Study on tissue culture is carried out to tail alpine ash DH32-13 kind, for the Fast-propagation of this non-defective unit kind, meet market supply, the ecological benefits and the economic benefit that improve these improved seeds are significant.
Summary of the invention
The object of the invention is for above deficiency, provide that a kind of operation easier is low, reproduction coefficient is high, the control cycle is short, can genetic stability and the tissue culture and rapid propagation method of the high tail alpine ash DH32-13 kind of survival rate.
Technical scheme of the present invention is as follows:
The tissue culture and rapid propagation method of tail alpine ash DH32-13 kind, concrete steps are as follows:
1. the foundation of explant: the DH32-13 kind through selecting, plant into pilot forest, February, select the age of tree of 4-5, robust growth, trunk be thick, without the select tree of damage by disease and insect, cut down from overhead 20-25 centimetre, after one month, stub starts to grow rudiment bar, waits that budling after 2 weeks is about 10 centimeters, semi-lignified, eye be when also sprouting, and adopts back budling as explant material.For reducing material contamination source, when taking budling, should select within continuous more than three days, carry out after fair weather.After the budling adopted back is removed most of blade, use tap water 15min, 0.5% washing powder solution soaks 15min, cleaner with tap water.
2. explant sterilization: the explant that 1. step obtains with 75% alcohol-pickled 30S, aseptic water washing 4-5 time, drains the water, then the mercuric chloride solution being placed in 0.1% is sterilized 9min, aseptic water washing 4-5 time, cut rudiment bar terminal bud get off second section rise 1-2 save stem with bud, long 1-2cm.
3. Initial culture: stem section step 2. processed aseptically is inoculated into Initial culture base, and Initial culture based component is: MS+NAA0.5mg/L+6-BA1.6mg/L+8% molasses, medium pH 5.9; Intensity of illumination 1000Lux, cultivation temperature 25 ± 2 DEG C, incubation time 35 ± 2 days.
4. squamous subculture: Initial culture goes out clump bud to joint director, and get clump bud and carry out squamous subculture, squamous subculture based component is: MS+ 6-BA 0.6mg/L+IAA0.25mg/L+0.15% amino-oligosaccharide+5% molasses; Cultivation temperature 25 ± 2 DEG C, light application time 16h/d, medium pH 5.8-6.0; Cultivate 20 days, form the budling with 3 stipes that 2cm is long, be cut into the segment of band stipes, each bud is divided into 2 sections; Every 20 days subcultures like this once, expand numerous test-tube plantlet.
5. culture of rootage: 4. step expands numerous test-tube plantlet and take out, and proceed to culture of rootage, culture of rootage based component is: 1/5MS+IBA0.5mg/L+1.0% amino-oligosaccharide+25% molasses; Intensity of illumination is 3000Lux, cultivation temperature 25 ± 2 DEG C, and be cultured to 25 days, root grows to 1-3cm, seedling grow tall 3cm time bottle outlet.
In subculture medium and root media, the Main Function of amino-oligosaccharide is: (1) excites clump bud tissue to produce phenolic compound, lignin and pathogenesis-related proteins, improves clump bud anti-adversity ability; (2) plant cell is activated, Promoting plant growth, the expression of regulating plant resistant gene, use amino-oligosaccharide as subculture medium and root media, inhibitory action can not be produced to the cultivation of explant under described dosage, in incubation, not be vulnerable to living contaminants, through multiple authentication, inventor proves that amino-oligosaccharide plays an important role in subculture medium of the present invention and root media, be its important component part.
And in three kinds of medium, add molasses (it is non crystallized containing sugar liquors that sugar refinery sugar industry is separated) also play an important role: (1), as the carbon source of tissue rapid propagation medium, substitutes sucrose, reduces production cost; (2) be easier to the various monose, thick protein and the mineral matter that utilize containing a large amount of plant, comparatively sucrose nutrition is more comprehensive.
Other medium compound can the textbooks of chemically dictionary or arviculture to its title and effect, the present invention adopts these compounds to carry out combining tail alpine ash DH32-13 kind the most applicable, particularly add the molasses being rich in nutrient component, be through that a lot of screening just finally determines.
6. hardening: select test-tube plantlet bottle outlet that is healthy and strong, semi-lignified to transplant at peat soil: coconut palm chaff proportioning is in the nutrient matrix of 1:3, keeps air humidity 85-90%, illumination 10000lux.
7. transplant; Hardening moved on to outdoor cultivation after 15 days, when seedling height in seedling-cultivating tray is 15-20cm, transplanted afforestation.
The tissue culture and rapid propagation method of tail alpine ash DH32-13 kind, also comprises the management after plantlet in vitro transplanting, is specially: before transplanting, plantlet in vitro is soaked 1-2h in normal root water; Use sufficient base manure, topdress in time, and prevention and elimination of disease and pests in time.
Described normal root water consists of: 3% metalaxyl+3% dislikes mould spirit+0.05% indolebutyric acid+0.05% Nafusaku+0.5% Sodium Polyacrylate+pure water surplus.
Described prevention and elimination of disease and pests is: prevent and treat damping off, fusarium wilt in seedling stage; Gray mold, leaf spot, defoliation is prevented and treated in vegetative period; Use bait formulation control termites, grub, cutworm; Thermal fog is used to prevent and treat eucalyptus caterpillar, looper in vegetative period.
Beneficial effect of the present invention is:
One, the merit of DH32-13 is maintained: fast growing, rotation short (industrial fiber material); Form is logical directly, branch is little, forest form is neat; Material is good, and texture is straight, barren-resistant, is applicable to large area region plantation.
Two, Eucalyptus Tissue Cultured quick-breeding method operation easier of the present invention is low, production cost is low, is easy to promote the use of on a large scale.In extensive tissue culture procedures, medium uses molasses as important component, instead of molasses, directly reduces the cost of medium; Meanwhile, in medium, amino-oligosaccharide appropriate use in the medium effectively inhibits miscellaneous bacteria to infect, and minimizing group training risk is the tissue culture and rapid propagation method of comparatively mature and reliable safety.
Three, the plantlet in vitro survival rate utilizing Eucalyptus Tissue Cultured quick-breeding method of the present invention to produce is high, and DH32-13 plantlet in vitro survival rate, more than 93%, is adapted at Guangxi most area plantation simultaneously and promotes, directly enhance economic benefit and social benefit.
Embodiment
Below preferred embodiment of the present invention is described in detail, can be easier to make advantages and features of the invention be readily appreciated by one skilled in the art, thus more explicit defining is made to protection scope of the present invention.
embodiment 1
The tissue culture and rapid propagation method of tail alpine ash DH32-13 kind, concrete steps are as follows:
1. the foundation of explant: the DH32-13 kind through selecting, plant into pilot forest, February, select the age of tree of 4-5, robust growth, trunk be thick, without the select tree of damage by disease and insect, cut down from overhead 20-25 centimetre, after one month, stub starts to grow rudiment bar, waits that budling after 2 weeks is about 10 centimeters, semi-lignified, eye be when also sprouting, and adopts back budling as explant material.For reducing material contamination source, when taking budling, should select within continuous more than three days, carry out after fair weather.After the budling adopted back is removed most of blade, use tap water 15min, 0.5% washing powder solution soaks 15min, cleaner with tap water.
2. explant sterilization: the explant that 1. step obtains with 75% alcohol-pickled 30S, aseptic water washing 4 times, drains the water, then the mercuric chloride solution being placed in 0.1% is sterilized 9min, aseptic water washing 5 times, cut rudiment bar terminal bud get off second section rise 1-2 save stem with bud, long 1-2cm.
3. Initial culture: stem section step 2. processed aseptically is inoculated into Initial culture base, and Initial culture based component is: MS+NAA0.5mg/L+6-BA1.6mg/L+8% molasses, medium pH 5.9; Intensity of illumination 1000Lux, cultivation temperature 25 ± 2 DEG C, incubation time 35 ± 2 days.
4. squamous subculture: Initial culture goes out clump bud to joint director, and get clump bud and carry out squamous subculture, squamous subculture based component is: MS+ 6-BA 0.6mg/L+IAA0.25mg/L+0.15% amino-oligosaccharide+5% molasses; Cultivation temperature 25 ± 2 DEG C, light application time 16h/d, medium pH 5.8-6.0; Cultivate 20 days, form the budling with 3 stipes that 2cm is long, be cut into the segment of band stipes, each bud is divided into 2 sections; Every 20 days subcultures like this once, expand numerous test-tube plantlet.
5. culture of rootage: treat that 4. step expands numerous test-tube plantlet and take out, proceed to culture of rootage, culture of rootage based component is: 1/5MS+IBA0.5mg/L+1.0% amino-oligosaccharide+25% molasses; Intensity of illumination is 3000Lux, cultivation temperature 25 ± 2 DEG C, and be cultured to 25 days, root grows to 1-3cm, seedling grow tall 3cm time bottle outlet.
6. hardening: select test-tube plantlet bottle outlet that is healthy and strong, semi-lignified to transplant at peat soil: coconut palm chaff proportioning is in the nutrient matrix of 1:3, keeps air humidity 85-90%, illumination 10000lux.
7. transplant; Hardening moved on to outdoor cultivation after 15 days, when seedling height in seedling-cultivating tray is 15-20cm, transplanted afforestation.
embodiment 2
The tissue culture and rapid propagation method of tail alpine ash DH32-13 kind, concrete steps are as follows:
1. the foundation of explant: the DH32-13 kind through selecting, plant into pilot forest, February, select the age of tree of 4-5, robust growth, trunk be thick, without the select tree of damage by disease and insect, cut down from overhead 20-25 centimetre, after one month, stub starts to grow rudiment bar, waits that budling after 2 weeks is about 10 centimeters, semi-lignified, eye be when also sprouting, and adopts back budling as explant material.For reducing material contamination source, when taking budling, should select within continuous more than three days, carry out after fair weather.After the budling adopted back is removed most of blade, use tap water 15min, 0.5% washing powder solution soaks 15min, cleaner with tap water.
2. explant sterilization: the explant that 1. step obtains with 75% alcohol-pickled 30S, aseptic water washing 5 times, drains the water, then the mercuric chloride solution being placed in 0.1% is sterilized 9min, aseptic water washing 4 times, cut rudiment bar terminal bud get off second section rise 1-2 save stem with bud, long 1-2cm.
3. Initial culture: stem section step 2. processed aseptically is inoculated into Initial culture base, and Initial culture based component is: MS+NAA0.5mg/L+6-BA1.6mg/L+8% molasses, medium pH 5.9; Intensity of illumination 1000Lux, cultivation temperature 25 ± 2 DEG C, incubation time 35 ± 2 days.
4. squamous subculture: Initial culture goes out clump bud to joint director, and get clump bud and carry out squamous subculture, squamous subculture based component is: MS+ 6-BA 0.6mg/L+IAA0.25mg/L+0.15% amino-oligosaccharide+5% molasses; Cultivation temperature 25 ± 2 DEG C, light application time 16h/d, medium pH 5.8-6.0; Cultivate 20 days, form the budling with 3 stipes that 2cm is long, be cut into the segment of band stipes, each bud is divided into 2 sections; Every 20 days subcultures like this once, expand numerous test-tube plantlet.
5. culture of rootage: treat that 4. step expands numerous test-tube plantlet and take out, proceed to culture of rootage, culture of rootage based component is: 1/5MS+IBA0.5mg/L+1.0% amino-oligosaccharide+25% molasses; Intensity of illumination is 3000Lux, cultivation temperature 25 ± 2 DEG C, and be cultured to 25 days, root grows to 1-3cm, seedling grow tall 3cm time bottle outlet.
6. hardening: select test-tube plantlet bottle outlet that is healthy and strong, semi-lignified to transplant at peat soil: coconut palm chaff proportioning is in the nutrient matrix of 1:3, keeps air humidity 85-90%, illumination 10000lux.
7. transplant; Hardening moved on to outdoor cultivation after 15 days, when seedling height in seedling-cultivating tray is 15-20cm, transplanted afforestation.
8. the management after transplanting: before transplanting, plantlet in vitro is soaked 1-2h in normal root water, normal root water consists of: 3% metalaxyl+3% dislikes mould spirit+0.05% indolebutyric acid+0.05% Nafusaku+0.5% Sodium Polyacrylate+pure water surplus; Every plantlet in vitro uses 10kg high-quality well-rotted farmyard manure, additional 2kgNPK15:15:15 composite fertilizer; Within latter 3 months, 6 months, respectively topdress 1 time in transplanting, topdress at every turn and use urea 1kg/, topdress every year once later, each use 2kg urea/, and prevention and elimination of disease and pests in time.Prevention and elimination of disease and pests and make with medicament as follows in DH32-13 process of growth: seedling blight and fusarium wilt only have fragmentary generation, after using normal root water to dilute 800 times to the plantlet in vitro that disease occurs, every plantlet in vitro perfusion water 500-800 gram; Gray mold uses the 5000 times of sprayings of BASF medicament 50% Boscalid water-dispersible grain dilution agent; Leaf spot and defoliation use 25% pyraclostrobin missible oil of BASF, dilute 8000 times of even spraying, and the effectively pre-disease prevention of decapacitation meets accident, and also has and obviously promotes Eucalyptus effect; Use 0.5% fipronil bait control soil insect grub cutworm and termite, using method is for be evenly sprinkling upon bait formulation around plantlet in vitro, and every uses bait formulation 20 grams-30 grams; When 3-5 age period eucalyptus outburst looper and caterpillar, the 1.8% A Wei chlorine cyanogen thermal fog control using Guangxi Tianyuan Biochemical Co., Ltd. to produce, saves man-hour, improves work efficiency, ensure that the high yield of eucalyptus.
Be below tail alpine ash DH32-13 plantlet in vitro breeding results that the inventive method obtains:
Can find out, tail alpine ash DH32-13 kind tissue culture and rapid propagation method of the present invention, reproduction coefficient is high, rooting rate reaches more than 99%, and survival rate is more than 97%, and plantlet in vitro can adapt to open-air atmosphere rapidly after transplanting, growth rapidly, anti-extraneous poor environment ability is strong, has planted popularization 366 hectares, has created larger economic benefit, ecological benefits and social benefit.
Claims (4)
1. a tissue culture and rapid propagation method for tail alpine ash DH32-13 kind, is characterized in that: comprise the following steps:
1. the foundation of explant: select the 4-5 age of tree, robust growth, trunk be thick, without the select tree of damage by disease and insect, cuts down from overhead 20-25cm, grows to 10cm, semi-lignified, eye be not when also sprouting, adopted back by budling as explant material until stub rudiment; For reducing material contamination source, when taking budling, should select within continuous more than three days, carry out after fair weather; After the budling adopted back is removed most of blade, use tap water 15min, 0.5% washing powder solution soaks 15min, cleaner with tap water;
2. explant sterilization: the explant that 1. step obtains with 75% alcohol-pickled 30S, aseptic water washing 4-5 time, drains the water, then the mercuric chloride solution being placed in 0.1% is sterilized 9min, aseptic water washing 4-5 time, cut rudiment bar terminal bud get off second section rise 1-2 joint stem with bud, long 1-2cm;
3. Initial culture: stem section step 2. processed aseptically is inoculated into Initial culture base, and Initial culture based component is: MS+NAA0.5mg/L+6-BA1.6mg/L+8% molasses, medium pH 5.9; Intensity of illumination 1000Lux, cultivation temperature 25 ± 2 DEG C, incubation time 35 ± 2 days;
4. squamous subculture: Initial culture goes out clump bud to joint director, and get clump bud and carry out squamous subculture, squamous subculture based component is: MS+ 6-BA 0.6mg/L+IAA0.25mg/L+0.15% amino-oligosaccharide+3% molasses; Cultivation temperature 25 ± 2 DEG C, light application time 16h/d, medium pH 5.8-6.0; Cultivate 20 days, form the budling with 3 stipes that 2cm is long, be cut into the segment of band stipes, each bud is divided into 2 sections; Every 20 days subcultures like this once, expand numerous test-tube plantlet;
5. culture of rootage: 4. step expands numerous test-tube plantlet and take out, and proceed to culture of rootage, culture of rootage based component is: 1/5MS+IBA0.5mg/L+1.0% amino-oligosaccharide+15% molasses; Intensity of illumination is 3000Lux, cultivation temperature 25 ± 2 DEG C, and be cultured to 25 days, root grows to 1-3cm, seedling grow tall 3cm time bottle outlet;
6. hardening: select test-tube plantlet bottle outlet that is healthy and strong, semi-lignified to transplant at peat soil: coconut palm chaff proportioning is in the nutrient matrix of 1:3, keeps air humidity 85-90%, illumination 10000lux;
7. transplant: hardening moved on to outdoor cultivation after 15 days, when seedling height in seedling-cultivating tray is 15-20cm, transplant afforestation.
2. the tissue culture and rapid propagation method of tail alpine ash DH32-13 kind according to claim 1, is characterized in that: also comprise the management after plantlet in vitro transplanting, be specially: before transplanting, plantlet in vitro is soaked 1-2h in normal root water; Use sufficient base manure, topdress in time, and prevention and elimination of disease and pests in time.
3. the tissue culture and rapid propagation method of tail alpine ash DH32-13 kind according to claim 2, is characterized in that: described normal root water consists of: 3% metalaxyl+3% dislikes mould spirit+0.05% indolebutyric acid+0.05% Nafusaku+0.5% Sodium Polyacrylate+pure water surplus.
4. the tissue culture and rapid propagation method of tail alpine ash DH32-13 kind according to claim 2, is characterized in that: described prevention and elimination of disease and pests is: prevent and treat damping off, fusarium wilt in seedling stage; Gray mold, leaf spot, defoliation is prevented and treated in vegetative period; Use bait formulation control termites, grub, cutworm; Thermal fog is used to prevent and treat eucalyptus caterpillar, looper in vegetative period.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201410590691.5A CN104285815B (en) | 2014-10-29 | 2014-10-29 | A kind of tissue culture and rapid propagation method of tail alpine ash DH32-13 kind |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201410590691.5A CN104285815B (en) | 2014-10-29 | 2014-10-29 | A kind of tissue culture and rapid propagation method of tail alpine ash DH32-13 kind |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN104285815A true CN104285815A (en) | 2015-01-21 |
| CN104285815B CN104285815B (en) | 2016-04-06 |
Family
ID=52305989
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201410590691.5A Active CN104285815B (en) | 2014-10-29 | 2014-10-29 | A kind of tissue culture and rapid propagation method of tail alpine ash DH32-13 kind |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN104285815B (en) |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105379624A (en) * | 2015-12-10 | 2016-03-09 | 广西壮族自治区国有东门林场 | Tissue culture fast propagation method of Eucalyptus pellita |
| CN107258536A (en) * | 2017-06-27 | 2017-10-20 | 广西壮族自治区国有东门林场 | A kind of eucalyptus breeding tail eucalyptus camaldulensis DH191 7 tissue culture and rapid propagation method |
| CN114586684A (en) * | 2022-01-26 | 2022-06-07 | 广西壮族自治区国有东门林场 | Tissue culture rapid propagation method of triploid eucalyptus new variety' Jinggui eucalyptus I |
| NL2035132B1 (en) * | 2023-02-10 | 2024-01-23 | Res Institute Of Tropical Forestry Chinese Academy Of Forestry | GENETIC TRANSFORMATION METHOD OF E. UROPHYLLA x E. GRANDIS DH3213 |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101884301A (en) * | 2010-04-08 | 2010-11-17 | 浙江省农业科学院 | Method for preventing pollution in plant tissue culture |
-
2014
- 2014-10-29 CN CN201410590691.5A patent/CN104285815B/en active Active
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101884301A (en) * | 2010-04-08 | 2010-11-17 | 浙江省农业科学院 | Method for preventing pollution in plant tissue culture |
Non-Patent Citations (4)
| Title |
|---|
| 刘奕清: "‘T13’号桉树再生体系的建立", 《中国种业》 * |
| 吴建宇等: "尾巨桉提纯复壮初步研究", 《现代农业科技》 * |
| 王笑春等: "桉树组织培养与再生系统建立研究", 《河北建筑科技学院学报》 * |
| 钱子刚: "《药用植物组织培养》", 31 January 2007, 中国中医药出版社 * |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105379624A (en) * | 2015-12-10 | 2016-03-09 | 广西壮族自治区国有东门林场 | Tissue culture fast propagation method of Eucalyptus pellita |
| CN107258536A (en) * | 2017-06-27 | 2017-10-20 | 广西壮族自治区国有东门林场 | A kind of eucalyptus breeding tail eucalyptus camaldulensis DH191 7 tissue culture and rapid propagation method |
| CN114586684A (en) * | 2022-01-26 | 2022-06-07 | 广西壮族自治区国有东门林场 | Tissue culture rapid propagation method of triploid eucalyptus new variety' Jinggui eucalyptus I |
| NL2035132B1 (en) * | 2023-02-10 | 2024-01-23 | Res Institute Of Tropical Forestry Chinese Academy Of Forestry | GENETIC TRANSFORMATION METHOD OF E. UROPHYLLA x E. GRANDIS DH3213 |
Also Published As
| Publication number | Publication date |
|---|---|
| CN104285815B (en) | 2016-04-06 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN104273037B (en) | A kind of tissue culture and rapid propagation method of tail alpine ash DH33-27 kind | |
| CN104285814B (en) | A kind of tissue culture and rapid propagation method of tail alpine ash DH32-43 kind | |
| CN101401524B (en) | Cutting propagation method for beach plum epicormic branch | |
| CN104365479B (en) | A kind of tissue culture and rapid propagation method of tail alpine ash DH32-28 kind | |
| CN103461121B (en) | Ex-vitro rooting method for tissue culture seedlings of pinus massoniana | |
| CN104686338A (en) | In-vitro culture technique of anther of angelica dahurica | |
| CN107018896B (en) | A kind of method of facility cuttage tilia miqueliana | |
| CN104285815B (en) | A kind of tissue culture and rapid propagation method of tail alpine ash DH32-13 kind | |
| CN109349105A (en) | A kind of iris tissue-cultured seedling mating system | |
| CN102415339A (en) | Rapid propagation method of photinia fraseri | |
| CN104304028B (en) | A kind of tissue culture and rapid propagation method of tail alpine ash DH32-26 kind | |
| CN104094848B (en) | The method of the induction of tung oil tree hypocotyledonery axis callus and highly efficient regeneration plant | |
| CN105941155A (en) | Method for rapidly propagating fraxinus mandshurica by utilizing suspension culture technology | |
| CN103299909A (en) | Method for breeding cherry seedlings in large scales through tissue culturing | |
| CN104823790B (en) | A kind of bulla walnut cuttage fast seedling-cultivating method | |
| CN104303765B (en) | The high-yield planting method of the stem of noble dendrobium | |
| CN102668991B (en) | Application of penicillin to simple test-tube breeding of grapes and novel technology for test-tube breeding of grapes | |
| CN104304026B (en) | A kind of tissue culture and rapid propagation method of tail alpine ash DH30-1 kind | |
| CN105638482B (en) | The method of walnut and Juglans mandshurica interspecific hybridization IMMATURE EMBRYOS CULTURE | |
| CN104304027B (en) | A kind of tissue culture and rapid propagation method of tail alpine ash DH32-29 kind | |
| CN104054579B (en) | A kind of method of tung oil tree petiole directly regenerated plant | |
| CN108450328A (en) | A kind of crocodile mouth flower quick breeding method for tissue culture | |
| CN103004600A (en) | Regeneration method for rosa wichuriana plant taking leaf as explant | |
| CN102119664B (en) | Method for culturing muscari botryoides mill with plant tissues | |
| CN106937601B (en) | A method of improving grape test tube seedling transplanting survival rate |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| TR01 | Transfer of patent right |
Effective date of registration: 20200826 Address after: 530000 Guangxi forest seedling demonstration base, Santang Town, Xingning District, Nanning City, Guangxi Zhuang Autonomous Region Patentee after: Guangxi Bagui forest flower seedlings Co.,Ltd. Address before: 532108, Dongmen County, Dongmen Town, Fusui County, the Guangxi Zhuang Autonomous Region, Chongzuo Patentee before: GUANGXI ZHUANG AUTONOMOUS REGION STATE-OWNED DONGMEN FOREST FARM |
|
| TR01 | Transfer of patent right | ||
| CP03 | Change of name, title or address |
Address after: No. 8, West Jiu Road, Keyuan, Xixiangtang District, Nanning City, Guangxi Zhuang Autonomous Region, 530007 Patentee after: Guangxi Bagui Seedling Hi-Tech Group Co., Ltd. Address before: Guangxi Forest Tree Seedling Demonstration Base, Santang Town, Xingning District, Nanning City, Guangxi Zhuang Autonomous Region, 530000 Patentee before: Guangxi Bagui forest flower seedlings Co.,Ltd. |
|
| CP03 | Change of name, title or address |