CN108522289A - A kind of Bougainvillea spectabilis detoxification method for tissue culture - Google Patents

A kind of Bougainvillea spectabilis detoxification method for tissue culture Download PDF

Info

Publication number
CN108522289A
CN108522289A CN201810661263.5A CN201810661263A CN108522289A CN 108522289 A CN108522289 A CN 108522289A CN 201810661263 A CN201810661263 A CN 201810661263A CN 108522289 A CN108522289 A CN 108522289A
Authority
CN
China
Prior art keywords
bougainvillea spectabilis
culture medium
culture
illumination
bougainvillea
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.)
Pending
Application number
CN201810661263.5A
Other languages
Chinese (zh)
Inventor
金红梅
李亚
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Ji'nan Yon Biological Technology Co Ltd
Original Assignee
Ji'nan Yon Biological Technology Co Ltd
Priority date (The priority date 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 date listed.)
Filing date
Publication date
Application filed by Ji'nan Yon Biological Technology Co Ltd filed Critical Ji'nan Yon Biological Technology Co Ltd
Priority to CN201810661263.5A priority Critical patent/CN108522289A/en
Publication of CN108522289A publication Critical patent/CN108522289A/en
Pending legal-status Critical Current

Links

Classifications

    • AHUMAN NECESSITIES
    • A01AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01HNEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
    • A01H4/00Plant reproduction by tissue culture techniques ; Tissue culture techniques therefor
    • A01H4/008Methods for regeneration to complete plants
    • AHUMAN NECESSITIES
    • A01AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01HNEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
    • A01H4/00Plant reproduction by tissue culture techniques ; Tissue culture techniques therefor
    • A01H4/001Culture apparatus for tissue culture

Landscapes

  • Life Sciences & Earth Sciences (AREA)
  • Developmental Biology & Embryology (AREA)
  • Engineering & Computer Science (AREA)
  • Biotechnology (AREA)
  • Cell Biology (AREA)
  • Botany (AREA)
  • Environmental Sciences (AREA)
  • Micro-Organisms Or Cultivation Processes Thereof (AREA)

Abstract

The invention discloses a kind of Bougainvillea spectabilis detoxification method for tissue culture, include the following steps:(1)Selection:Training choose sprout within 1 ~ 2 year it is full, without rotten disease-free, healthy and strong bougainvillea spectabilis branch;(2)Vernalization:The bougainvillea spectabilis branch chosen is subjected to vernalization;(3)Surface treatment;(4)Disinfection;(5)Cutting;(6)Initial culture;(7)Proliferation and subculture:Aseptic seedling is inoculated into progress proliferation and subculture culture on corresponding culture medium;(8)Culture of rootage;(9)Tame hardening.The beneficial effects of the invention are as follows:By tissue cultures Bougainvillea spectabilis quality and quality is greatly improved, and reduce cost to greatest extent in Bougainvillea spectabilis, improves breeding coefficient, and providing technical basis for Bougainvillea spectabilis test tube seedling factorial praluction is of great significance;The technical barrier in Bougainvillea spectabilis tissue culture procedures has been captured, good culture effect is reached.

Description

A kind of Bougainvillea spectabilis detoxification method for tissue culture
Technical field
The invention belongs to field of plant tissue culture technique, more particularly to a kind of Bougainvillea spectabilis detoxification tissue cultures side Method.
Background technology
Bougainvillea spectabilis is the evergreen wooden big rattan wood plant of Nyctaginaceae bougainvillea, belongs to the sight flowering shrub class in flowering tree, It is important ornamental flower and excellent garden plants.It is had a graceful carriage, the light green soft cottonrose hibiscus of branches and leaves, from the spring in early autumn to next year End, flowering time is for half a year, and the flowers extensive as a kind of Unique, cultivation management are very popular.Triangle When plum is as flower culture, it is often trimmed to shrub, dungarunga shape or multilayer pagoda shape, therefore is cried " paper flower " in Japan;Again because of it Colored bract is lobate, ellipse or ellipticity lanceolar, there is the multiple colors such as red, white, orange, purple, secondary color, rich in variety like Du Cuckoo, therefore be referred to as " strangling cuckoo " in Guangzhou;Again since its shoot top is typically three fasciations among three pieces bract, flower The beautiful purple goblin of bract of periphery is red, is similar to triangle, therefore in Fujian also known as " Bougainvillea spectabilis " or " bougainvillea "." precious towel is in botany Transliteration of the flower " then from its Classification system.It may then be gained the name due to it blooming around the Spring Festival " gathering red " in Hong Kong.
Bougainvillea spectabilis platymiscium serves not only as flowers and is concerned, and the research in recent years in relation to Bougainvillea spectabilis other application value makes people There is new understanding to it.The isolated pine camphor with para-insulin function is to substituting insulin therapy from Bougainvillea spectabilis blade The side effect that diabetes and mitigation insulinization are brought is of great significance.Research is it is also shown that in Bougainvillea spectabilis blade and root At least contain 2 kinds of antiviral protein-BAPI and Bouganin, they have different degrees of suppression for various plants virus It makes and uses.These results of study have very positive meaning for the efficient biological pesticide of exploitation low toxicity.Bougainvillea spectabilis is still ground Study carefully one of the important model plant of beet pigments metabolism, the type and content of beet pigments are enriched for the day of exploitation safety Right food coloring opens wide foreground.These correlative studys illustrate multi-value of the Bougainvillea spectabilis as resource plant, are Further the ornamental of exploitation Bougainvillea spectabilis, medicine, biological pesticide are worth and make as natural food colour and performance environment remediation With providing sufficient scientific basis.
In recent years, it is domestic that be concentrated mainly on biological characteristics, cultivation management, breeding, cuttage for the research of Bougainvillea spectabilis numerous It grows, Greenland Application etc..As people are to deepening continuously of studying of Bougainvillea spectabilis and the fast development of Urban Landscape Construction cause, The traditional modes of reproduction of Bougainvillea spectabilis cannot increasingly meet the needs of market, because there are reproduction speeds for traditional modes of reproduction Slowly, the drawbacks such as growth coefficient is small, and rootage duration is long, repoductive time is restricted;And with Bougainvillea spectabilis breeding research work into Exhibition, people have cultivated the higher new varieties of many ornamental values, some new varieties are relatively more tired with traditional propagation method breeding It is difficult, it is difficult to the nursery stock that a large amount of improved seeds or strain are provided in a short time, to affect expanding propagation and the city of Bougainvillea spectabilis Field possession share.
Invention content
The purpose of the present invention is to overcome the defect of above-mentioned Bougainvillea spectabilis tissue cultures, to the detoxification tissue cultures side of Bougainvillea spectabilis Method carries out systematic research, by many experiments, obtains a kind of culture side easy to operate, improving Bougainvillea spectabilis yield and quality Method.
To achieve the above object, the present invention uses following technical proposals:
A kind of Bougainvillea spectabilis detoxification method for tissue culture, including:
(1) selection:Training chooses life in 1 ~ 2 year without rotten disease-free, healthy and strong bougainvillea spectabilis branch;
(2) vernalization:The bougainvillea spectabilis branch chosen is inserted in the bottle with water, is placed in illumination room, 25 ~ 30 DEG C of temperature is maintained, 3000 ~ 5000Lux of intensity of illumination, light application time for 24 hours, irradiate 5 ~ 10d, choose 3 ~ 7cm Bougainvillea spectabilis sprout segments;
(3) it is surface-treated:The Bougainvillea spectabilis sprout segment gathered is rinsed well using flowing water, 10 are cleaned with 2% Tween-80 ~ 15min is rinsed well using flowing water;
(4) it sterilizes:It is impregnated using 75% alcohol, 0.1%HgCl 5 ~ 10min of solution disinfection, aseptic water washing is clean;
(5) it cuts:The Bougainvillea spectabilis sprout segment notch disinfected is updated processing, retains 2 ~ 3cm;
(6) Initial culture:The Bougainvillea spectabilis segment handled well is seeded on culture medium I, is positioned over 25 ~ 28 DEG C of temperature, per the daylight It is 1000 ~ 3000Lux according to 18 ~ 22h, intensity of illumination, 40 ~ 50d of culture;
(7) proliferation and subculture:Selecting step(6)In obtained aseptic seedling be inoculated on culture medium II and carry out Multiplying culture, train Foster condition is:25 ~ 28 DEG C of temperature, daily 10 ~ 14h of illumination, intensity of illumination are 5000 ~ 7000Lux, cultivate 25 ~ 35d;In repetition State breeding;
(8) it takes root:Choose growing way stalwartness Bougainvillea spectabilis seedling, be seeded in culture medium III, be positioned over 25 ~ 28 DEG C of temperature, often In 12 ~ 16h of its illumination, the environment that intensity of illumination is 5000 ~ 7000Lux, 20 ~ 30d is cultivated;
(9) hardening is tamed:The Bougainvillea spectabilis seedling for the well developed root system that step (8) obtains is transferred in small temperature canopy and is tamed, is shaded, It is 15 ~ 30 DEG C to keep temperature in small temperature canopy, and humid control humidity after 85 ~ 95%, 7d is decremented to 80%, hereafter gradually successively decreases, 15d Bougainvillea spectabilis seedling is removed from tissue culture medium (TCM) afterwards, the culture medium of root is cleaned up, in plantation to substrate soil;
Wherein:Culture medium I is 0.5 ~ 1.5mg/LIBA of addition, 1.0 ~ 3.0mg/LZT, 25 ~ 35g/L sucrose in MS improved culture mediums With 2 ~ 5g/L agar;
Culture medium II be MS improved culture mediums in addition 4 ~ 5mg/LZT, 0.01 ~ 0.5mg/LIAA, 0.1 ~ 1mg/LNAA, 25 ~ 35g/L sucrose and 2 ~ 5g/L agar;
Culture medium III be 0.1 ~ 1mg/LNAA of addition, 0.1 ~ 0.5mg/LIBA, 15 ~ 25g/L sucrose and 2 in MS improved culture mediums ~ 5g/L agar.
As an optimization, a kind of Bougainvillea spectabilis detoxification method for tissue culture, including:
(1) selection:Training chooses life in 1 ~ 2 year without rotten disease-free, healthy and strong bougainvillea spectabilis branch;
(2) vernalization:The bougainvillea spectabilis branch chosen is inserted in the bottle with water, is placed in illumination room, 25 ~ 30 DEG C of temperature is maintained, 4000 ~ 5000Lux of intensity of illumination, light application time for 24 hours, irradiate 7d, choose 4 ~ 6cm Bougainvillea spectabilis sprout segments;
(3) it is surface-treated:The Bougainvillea spectabilis sprout segment gathered is rinsed well using flowing water, 10 are cleaned with 2% Tween-80 ~ 15min is rinsed well using flowing water;
(4) it sterilizes:It is impregnated using 75% alcohol, 0.1%HgCl 5 ~ 10min of solution disinfection, aseptic water washing is clean;
(5) it cuts:The Bougainvillea spectabilis sprout segment notch disinfected is updated processing, retains 2 ~ 3cm;
(6) Initial culture:The Bougainvillea spectabilis segment handled well is seeded on culture medium I, is positioned over 25 ~ 26 DEG C of temperature, per the daylight According to 20h, intensity of illumination 2000Lux, 45 ~ 50d of culture;
(7) proliferation and subculture:Selecting step(6)In obtained aseptic seedling be inoculated on culture medium II and carry out Multiplying culture, train Foster condition is:25 ~ 26 DEG C of temperature, daily illumination 12h, intensity of illumination are 5000 ~ 7000Lux, cultivate 25 ~ 30d;Repeat above-mentioned increasing Grow process;
(8) it takes root:Choose growing way stalwartness Bougainvillea spectabilis aseptic seedling, be seeded in culture medium III, be positioned over 25 ~ 26 DEG C of temperature, In daily illumination 14h, the environment that intensity of illumination is 5000 ~ 7000Lux, 25 ~ 30d is cultivated;
(9) hardening is tamed:The Bougainvillea spectabilis seedling for the well developed root system that step (8) obtains is transferred in small temperature canopy and is tamed, is shaded, It is 20 ~ 30 DEG C to keep temperature in small temperature canopy, and humid control humidity after 85 ~ 95%, 7d is decremented to 80%, hereafter gradually successively decreases, 15d Bougainvillea spectabilis seedling is removed from tissue culture medium (TCM) afterwards, the culture medium of root is cleaned up, in plantation to substrate soil;
As an optimization, culture medium I be MS improved culture mediums in addition 0.5 ~ 1.5mg/LIBA, 1.0 ~ 3.0mg/LZT, 25 ~ 35g/L sucrose and 2 ~ 5g/L agar.
As an optimization, culture medium II be MS improved culture mediums in addition 4 ~ 5mg/LZT, 0.01 ~ 0.5mg/LIAA, 0.1 ~ 1mg/LNAA, 25 ~ 35g/L sucrose and 2 ~ 5g/L agar.
As an optimization, culture medium III is 0.1 ~ 1mg/LNAA of addition, 0.1 ~ 0.5mg/LIBA, 15 in MS improved culture mediums ~ 25g/L sucrose and 2 ~ 5g/L agar.
As an optimization, the pH of the culture medium I, culture medium II and culture medium II is adjusted to 5.8.
As an optimization, the alcohol disinfecting time is no more than 1min in the step (4).
As an optimization, the step(9)Artificially feed is carried out under the conditions of the middle domestication hardening link cloudy day, light filling intensity is 3000~5000Lux。
As an optimization, the step(9)Middle domestication hardening step, be transferred to small temperature canopy after 10 ~ 15 days daily at dusk or Early morning divulges information half an hour.
It is tobacco cell Training Design that MS culture mediums, which were Murashige and Skoog in 1962, its main feature is that inorganic salts It is higher with ion concentration, it is more stable ionic equilibrium solution, its nitrate content is high, and the quantity and ratio of nutrient are closed It is suitable, the nutrition and physiological requirements of plant cell can be met, thus the scope of application is wider, most plants tissue-culturing quick-propagation Use it as the minimal medium of culture medium.MS improved culture mediums are to remove inositol on the basis of MS culture mediums, and ammonium nitrate subtracts Half, increase ascorbic acid 10mg/L.
Heteroauxin (IAA):The Endogenous auxin of generally existing in a kind of plant, belongs to Benzazole compounds, You Mingzhuo Long element, auxin, different growth hormone, are a kind of organic matters, are stablized to visible light, have duality, plant different plant growth Position is more than stem to its susceptibility difference, general root, and different plants are also different to its sensitivity.
Indolebutyric acid(IBA):Plant growth regulator is used for cell division and hyperplasia, promotes draft and woody plant The root of object it is mitogenetic.
Methyl α-naphthyl acetate (NAA):It is the intermediate for the plant growth regulator and naphthaleneacetamide for promoting plant root growth. Methyl α-naphthyl acetate is used as plant growth regulator, has and promotes cell division and expand, induced synthesis adventitious root, increase is beared fruit, prevented Shedding etc., can be with nutrition stream transporting to site of action.
Zeatin(ZT), it is a kind of basic element of cell division, belongs to six big natural phytohormone, that is, auxin, gibberellin, cell point One of element, abscisic acid, ethylene and brassin lactones are split, corn is named as because being separated to a kind of analog from corn for the first time Element, its physiological function is more complicated, including promotes plant cell division, prevents chlorophyll and protein degradation, slows down breathing Effect keeps cell viability, delays plant senesecence etc..
Agar:The most useful characteristic of agar is that the temperature difference between its condensation point and fusing point is very big.It needs to add in water Just start to melt when heat is to 95 DEG C, just start to solidify when the solution temperature after fusing need to drop to 40 DEG C, so it is to prepare solid The best coagulator of culture medium.The solid medium prepared with agar, is used for high-temperature cultivation and non-fusible, in solidification When preceding inoculation, culture will not also be scalded dead.Therefore, agar is to prepare most widely used one kind in various biological mediums to coagulate Gu agent.
Sucrose:Sucrose is in addition to energy supply, moreover it is possible to which evoked callus tissue breaks up again;When culture medium high-temperature sterilization, sugarcane Sugar is more stable and is not easy to be carbonized;Contain invertase in plant cell, sucrose can preferably remain hypotonic in culture medium Environment, the osmotic pressure that sucrose is formed can keep relative stability for a long time, can reduce the pollution of microorganism to a certain extent.
The beneficial effects of the invention are as follows:
1. Bougainvillea spectabilis can play the role of detoxification, fast numerous, Germ-plasma resources protection, quality-improving, significantly by tissue cultures Bougainvillea spectabilis quality and quality are improved, the planting scale of Bougainvillea spectabilis is expanded.
2. the medium component that control differentiation, subculture grow, take root etc. is determined in tissue cultures, growth has been determined The optimum optimization selection of the type, concentration and condition of culture of Auto-regulator, has captured the skill in Bougainvillea spectabilis tissue culture procedures Art barrier reaches good culture effect.
Description of the drawings
In order to more clearly explain the embodiment of the invention or the technical proposal in the existing technology, to embodiment or will show below There is attached drawing needed in technology description to be briefly described, it should be apparent that, the accompanying drawings in the following description is only this Some embodiments of invention for those of ordinary skill in the art without creative efforts, can be with Obtain other attached drawings according to these attached drawings.
Fig. 1 is the Initial culture step Bougainvillea spectabilis schematic diagram of the present invention;
Fig. 2 is proliferation and subculture step Bougainvillea spectabilis schematic diagram of the present invention;
Fig. 3 is that the present invention is taken root step Bougainvillea spectabilis schematic diagram.
Specific implementation mode
The following is a clear and complete description of the technical scheme in the embodiments of the invention, it is clear that described embodiment Only a part of the embodiment of the present invention, instead of all the embodiments.Based on the embodiments of the present invention, the common skill in this field The every other embodiment that art personnel are obtained without creative efforts belongs to the model that the present invention protects It encloses.In the following examples, the experimental methods for specific conditions are not specified, usually according to normal condition, or according to proposed by manufacturer Condition, unless stated otherwise, ratio and percentage are based on weight.
It is commercially available that the embodiment of the present invention is raw materials used, culture medium I in the embodiment, culture medium II and culture medium II PH is adjusted to 5.8.
Embodiment 1, a kind of Bougainvillea spectabilis detoxification method for tissue culture, including:
(1) selection:Training chooses life in 1 ~ 2 year without rotten disease-free, healthy and strong bougainvillea spectabilis branch;
(2) vernalization:The bougainvillea spectabilis branch chosen is inserted in the bottle with water, is placed in illumination room, 28 ~ 30 DEG C of temperature is maintained, Intensity of illumination 3000Lux, light application time for 24 hours, irradiate 7d, choose 3 ~ 7cm Bougainvillea spectabilis sprout segments;
(3) it is surface-treated:The Bougainvillea spectabilis sprout segment gathered is rinsed well using flowing water, 10 are cleaned with 2% Tween-80 ~ 15min is rinsed well using flowing water;
(4) it sterilizes:55s, 0.1%HgCl solution disinfection 5min are impregnated using 75% alcohol, aseptic water washing is clean;
(5) it cuts:The Bougainvillea spectabilis sprout segment notch disinfected is updated processing, retains 2 ~ 3cm;
(6) Initial culture:The Bougainvillea spectabilis segment handled well is seeded on culture medium I, is positioned over 26 ~ 27 DEG C of temperature, per the daylight According to 22h, intensity of illumination 1000Lux, culture 45d;Culture medium I is that 0.5mg/LIBA, 2.0mg/ are added in MS improved culture mediums LZT, 35g/L sucrose and 2g/L agar;
(7) proliferation and subculture:Selecting step(6)In obtained aseptic seedling be inoculated on culture medium II and carry out Multiplying culture, train Foster condition is:27 ~ 28 DEG C of temperature, daily illumination 10h, intensity of illumination 6000Lux, culture 35d;Repeat above-mentioned breeding; Culture medium II is addition 4.5mg/LZT, 0.5mg/LIAA, 0.1mg/LNAA, 30g/L sucrose and 5g/L fine jades in MS improved culture mediums Fat;
(8) it takes root:Choose growing way stalwartness Bougainvillea spectabilis aseptic seedling, be seeded in culture medium III, be positioned over 25 ~ 26 DEG C of temperature, In daily illumination 14h, the environment that intensity of illumination is 7000Lux, 20d is cultivated;Culture medium III is to be added in MS improved culture mediums 0.1mg/LNAA, 0.2mg/LIBA, 25g/L sucrose and 2g/L agar.The Bougainvillea spectabilis aseptic seedling of well developed root system, the item number of root is 7 ~ 8, length is 10 ~ 20min, is conveniently attached in substrate soil, avoids hindering root.
(9) hardening is tamed:The Bougainvillea spectabilis seedling for the well developed root system that step (8) obtains is transferred in small temperature canopy and is tamed, Shade, it is 20 ~ 25 DEG C to keep temperature in small temperature canopy, and humid control humidity after 85 ~ 95%, 7d is decremented to 80%, hereafter gradually passs Subtract, be transferred to small temperature canopy after 10 ~ 15 days daily at dusk or ventilation half an hour in early morning, by Bougainvillea spectabilis seedling from tissue culture medium (TCM) after 15d Middle removal cleans up the culture medium of root, in plantation to substrate soil;Artificially feed, light filling intensity are carried out under the conditions of cloudy day For 3000Lux.
The Bougainvillea spectabilis shoot survival percent obtained by this Bougainvillea spectabilis detoxification method for tissue culture is 99% or more, Bougainvillea spectabilis yield All it is greatly improved with quality.
A kind of 2 Bougainvillea spectabilis detoxification method for tissue culture of embodiment, including:
(1) selection:Training chooses life in 1 ~ 2 year without rotten disease-free, healthy and strong bougainvillea spectabilis branch;
(2) vernalization:The bougainvillea spectabilis branch chosen is inserted in the bottle with water, is placed in illumination room, 26 ~ 28 DEG C of temperature is maintained, Intensity of illumination 5000Lux, light application time for 24 hours, irradiate 5d, choose 3 ~ 7cm Bougainvillea spectabilis sprout segments;
(3) it is surface-treated:The Bougainvillea spectabilis sprout segment gathered is rinsed well using flowing water, 10 are cleaned with 2% Tween-80 ~ 15min is rinsed well using flowing water;
(4) it sterilizes:55s, 0.1%HgCl solution disinfection 10min are impregnated using 75% alcohol, aseptic water washing is clean;
(5) it cuts:The Bougainvillea spectabilis sprout segment notch disinfected is updated processing, retains 2 ~ 3cm;
(6) Initial culture:The Bougainvillea spectabilis segment handled well is seeded on culture medium I, is positioned over 25 ~ 26 DEG C of temperature, per the daylight According to 20h, intensity of illumination 3000Lux, culture 40d;Culture medium I is that 1.5mg/LIBA, 1.0mg/ are added in MS improved culture mediums LZT, 30g/L sucrose and 5g/L agar;
(7) proliferation and subculture:Selecting step(6)In obtained aseptic seedling be inoculated on culture medium II and carry out Multiplying culture, train Foster condition is:26 ~ 27 DEG C of temperature, daily illumination 14h, intensity of illumination 5000Lux, culture 30d;Repeat above-mentioned breeding; Culture medium II is addition 4mg/LZT, 0.3mg/LIAA, 1mg/LNAA, 25g/L sucrose and 3.5g/L fine jades in MS improved culture mediums Fat;
(8) it takes root:Choose growing way stalwartness Bougainvillea spectabilis aseptic seedling, be seeded in culture medium III, be positioned over 27 ~ 28 DEG C of temperature, In daily illumination 12h, the environment that intensity of illumination is 6000Lux, 30d is cultivated;Culture medium III is to be added in MS improved culture mediums 1mg/LNAA, 0.1mg/LIBA, 20g/L sucrose and 5g/L agar.The item number of the Bougainvillea spectabilis aseptic seedling of well developed root system, root is 7 ~ 8 Item, length are 10 ~ 20min, are conveniently attached in substrate soil, avoid hindering root.
(9) hardening is tamed:The Bougainvillea spectabilis seedling for the well developed root system that step (8) obtains is transferred in small temperature canopy and is tamed, Shade, it is 15 ~ 20 DEG C to keep temperature in small temperature canopy, and humid control humidity after 85 ~ 90%, 7d is decremented to 80%, hereafter gradually passs Subtract, be transferred to small temperature canopy after 10 ~ 15 days daily at dusk or ventilation half an hour in early morning, by Bougainvillea spectabilis seedling from tissue culture medium (TCM) after 15d Middle removal cleans up the culture medium of root, in plantation to substrate soil;Artificially feed, light filling intensity are carried out under the conditions of cloudy day For 4000Lux.
The Bougainvillea spectabilis shoot survival percent obtained by this Bougainvillea spectabilis detoxification method for tissue culture is 99% or more, Bougainvillea spectabilis yield All it is greatly improved with quality.
A kind of 3 Bougainvillea spectabilis detoxification method for tissue culture of embodiment, including:
(1) selection:Training chooses life in 1 ~ 2 year without rotten disease-free, healthy and strong bougainvillea spectabilis branch;
(2) vernalization:The bougainvillea spectabilis branch chosen is inserted in the bottle with water, is placed in illumination room, 25 ~ 26 DEG C of temperature is maintained, Intensity of illumination 4000Lux, light application time for 24 hours, irradiate 10d, choose 3 ~ 7cm Bougainvillea spectabilis sprout segments;
(3) it is surface-treated:The Bougainvillea spectabilis sprout segment gathered is rinsed well using flowing water, 10 are cleaned with 2% Tween-80 ~ 15min is rinsed well using flowing water;
(4) it sterilizes:55s, 0.1%HgCl solution disinfection 10min are impregnated using 75% alcohol, aseptic water washing is clean;
(5) it cuts:The Bougainvillea spectabilis sprout segment notch disinfected is updated processing, retains 2 ~ 3cm;
(6) Initial culture:The Bougainvillea spectabilis segment handled well is seeded on culture medium I, is positioned over 27 ~ 28 DEG C of temperature, per the daylight According to 18h, intensity of illumination 2000Lux, culture 50d;Culture medium I is that 1.0mg/LIBA, 3.0mg/ are added in MS improved culture mediums LZT, 25g/L sucrose and 3.5g/L agar;
(7) proliferation and subculture:Selecting step(6)In obtained aseptic seedling be inoculated on culture medium II and carry out Multiplying culture, train Foster condition is:25 ~ 26 DEG C of temperature, daily illumination 12h, intensity of illumination 7000Lux, culture 25d;Repeat above-mentioned breeding; Culture medium II is addition 5mg/LZT, 0.01mg/LIAA, 0.5mg/LNAA, 35g/L sucrose and 2g/L fine jades in MS improved culture mediums Fat;
(8) it takes root:Choose growing way stalwartness Bougainvillea spectabilis aseptic seedling, be seeded in culture medium III, be positioned over 26 ~ 27 DEG C of temperature, In daily illumination 16h, the environment that intensity of illumination is 5000Lux, 25d is cultivated;Culture medium III is to be added in MS improved culture mediums 0.5mg/LNAA, 0.5mg/LIBA, 15g/L sucrose and 3.5g/L agar.The item number of the Bougainvillea spectabilis aseptic seedling of well developed root system, root is 7 ~ 8, length is 10 ~ 20min, is conveniently attached in substrate soil, avoids hindering root.
(9) hardening is tamed:The Bougainvillea spectabilis seedling for the well developed root system that step (8) obtains is transferred in small temperature canopy and is tamed, Shade, it is 25 ~ 30 DEG C to keep temperature in small temperature canopy, and humid control humidity after 85 ~ 95%, 7d is decremented to 80%, hereafter gradually passs Subtract, be transferred to small temperature canopy after 10 ~ 15 days daily at dusk or ventilation half an hour in early morning, by Bougainvillea spectabilis seedling from tissue culture medium (TCM) after 15d Middle removal cleans up the culture medium of root, in plantation to substrate soil;Artificially feed, light filling intensity are carried out under the conditions of cloudy day For 3000Lux.
The Bougainvillea spectabilis shoot survival percent obtained by this Bougainvillea spectabilis detoxification method for tissue culture is 99% or more, Bougainvillea spectabilis yield All it is greatly improved with quality.
A kind of 4 Bougainvillea spectabilis detoxification method for tissue culture of embodiment, including:
(1) selection:Training chooses life in 1 ~ 2 year without rotten disease-free, healthy and strong bougainvillea spectabilis branch;
(2) vernalization:The bougainvillea spectabilis branch chosen is inserted in the bottle with water, is placed in illumination room, 25 ~ 30 DEG C of temperature is maintained, 4000 ~ 5000Lux of intensity of illumination, light application time for 24 hours, irradiate 7d, choose 4 ~ 6cm Bougainvillea spectabilis sprout segments;
(3) it is surface-treated:The Bougainvillea spectabilis sprout segment gathered is rinsed well using flowing water, 10 are cleaned with 2% Tween-80 ~ 15min is rinsed well using flowing water;
(4) it sterilizes:It is impregnated using 75% alcohol, 0.1%HgCl 5 ~ 10min of solution disinfection, aseptic water washing is clean;
(5) it cuts:The Bougainvillea spectabilis sprout segment notch disinfected is updated processing, retains 2 ~ 3cm;
(6) Initial culture:The Bougainvillea spectabilis segment handled well is seeded on culture medium I, is positioned over 25 ~ 26 DEG C of temperature, per the daylight According to 20h, intensity of illumination 2000Lux, culture 45d;Culture medium I is that 1.0mg/LIBA, 2.0mg/ are added in MS improved culture mediums LZT, 30g/L sucrose and 3.5g/L agar.
(7) proliferation and subculture:Selecting step(6)In obtained aseptic seedling be inoculated on culture medium II and carry out proliferation training It supports, condition of culture is:25 ~ 26 DEG C of temperature, daily illumination 12h, intensity of illumination are 5500 ~ 6000Lux, cultivate 28d;It repeats above-mentioned Breeding;Culture medium II is that 3mg/LZT, 0.1mg/LIAA, 0.5mg/LNAA, 30g/L sucrose are added in MS improved culture mediums With 3.5g/L agar.
(8) it takes root:Choose growing way stalwartness Bougainvillea spectabilis aseptic seedling, be seeded in culture medium III, be positioned over temperature 25 ~ 26 DEG C, daily illumination 14h, intensity of illumination be 6000 ~ 6500Lux environment in, cultivate 28d;Culture medium III is MS improvement cultures 0.5mg/LNAA, 0.2mg/LIBA, 20g/L sucrose and 3.5g/L agar are added in base.The Bougainvillea spectabilis aseptic seedling of well developed root system, root Item number be 7 ~ 8, length be 10 ~ 20min, be conveniently attached in substrate soil, avoid hindering root.
(9) hardening is tamed:The Bougainvillea spectabilis seedling for the well developed root system that step (8) obtains is transferred in small temperature canopy and is tamed, Shade, it is 20 ~ 28 DEG C to keep temperature in small temperature canopy, and humid control humidity after 85 ~ 95%, 7d is decremented to 80%, hereafter gradually passs Subtract, be transferred to small temperature canopy after 10 ~ 15 days daily at dusk or ventilation half an hour in early morning, by Bougainvillea spectabilis seedling from tissue culture medium (TCM) after 15d Middle removal cleans up the culture medium of root, in plantation to substrate soil;Artificially feed, light filling intensity are carried out under the conditions of cloudy day For 3500 ~ 4000Lux.
The Bougainvillea spectabilis shoot survival percent obtained by this Bougainvillea spectabilis detoxification method for tissue culture is 99% or more, Bougainvillea spectabilis yield All it is greatly improved with quality.
The foregoing is merely illustrative of the preferred embodiments of the present invention, is not intended to limit the invention, all essences in the present invention With within principle, any modification, equivalent replacement, improvement and so on should all be included in the protection scope of the present invention god.

Claims (9)

1. a kind of Bougainvillea spectabilis detoxification method for tissue culture, it is characterised in that:Including:
(1) selection:Training chooses life in 1 ~ 2 year without rotten disease-free, healthy and strong bougainvillea spectabilis branch;
(2) vernalization:The bougainvillea spectabilis branch chosen is inserted in the bottle with water, is placed in illumination room, 25 ~ 30 DEG C of temperature is maintained, 3000 ~ 5000Lux of intensity of illumination, light application time for 24 hours, irradiate 5 ~ 10d, choose 3 ~ 7cm Bougainvillea spectabilis sprout segments;
(3) it is surface-treated:The Bougainvillea spectabilis sprout segment gathered is rinsed well using flowing water, 10 are cleaned with 2% Tween-80 ~ 15min is rinsed well using flowing water;
(4) it sterilizes:It is impregnated using 75% alcohol, 0.1%HgCl 5 ~ 10min of solution disinfection, aseptic water washing is clean;
(5) it cuts:The Bougainvillea spectabilis sprout segment notch disinfected is updated processing, retains 2 ~ 3cm;
(6) Initial culture:The Bougainvillea spectabilis segment handled well is seeded on culture medium I, is positioned over 25 ~ 28 DEG C of temperature, per the daylight It is 1000 ~ 3000Lux according to 18 ~ 22h, intensity of illumination, 40 ~ 50d of culture;
(7) proliferation and subculture:Selecting step(6)In obtained aseptic seedling be inoculated on culture medium II and carry out Multiplying culture, train Foster condition is:25 ~ 28 DEG C of temperature, daily 10 ~ 14h of illumination, intensity of illumination are 5000 ~ 7000Lux, cultivate 25 ~ 35d;In repetition State breeding;
(8) it takes root:Choose growing way stalwartness Bougainvillea spectabilis seedling, be seeded in culture medium III, be positioned over 25 ~ 28 DEG C of temperature, often In 12 ~ 16h of its illumination, the environment that intensity of illumination is 5000 ~ 7000Lux, 20 ~ 30d is cultivated;
(9) hardening is tamed:The Bougainvillea spectabilis seedling for the well developed root system that step (8) obtains is transferred in small temperature canopy and is tamed, is shaded, It is 15 ~ 30 DEG C to keep temperature in small temperature canopy, and humid control humidity after 85 ~ 95%, 7d is decremented to 80%, hereafter gradually successively decreases, 15d Bougainvillea spectabilis seedling is removed from tissue culture medium (TCM) afterwards, the culture medium of root is cleaned up, in plantation to substrate soil;
Wherein:Culture medium I is 0.5 ~ 1.5mg/LIBA of addition, 1.0 ~ 3.0mg/LZT, 25 ~ 35g/L sucrose in MS improved culture mediums With 2 ~ 5g/L agar;
Culture medium II be MS improved culture mediums in addition 4 ~ 5mg/LZT, 0.01 ~ 0.5mg/LIAA, 0.1 ~ 1mg/LNAA, 25 ~ 35g/L sucrose and 2 ~ 5g/L agar;
Culture medium III be 0.1 ~ 1mg/LNAA of addition, 0.1 ~ 0.5mg/LIBA, 15 ~ 25g/L sucrose and 2 in MS improved culture mediums ~ 5g/L agar.
2. a kind of Bougainvillea spectabilis detoxification method for tissue culture according to claim 1, it is characterised in that:Including:
(1) selection:Training chooses life in 1 ~ 2 year without rotten disease-free, healthy and strong bougainvillea spectabilis branch;
(2) vernalization:The bougainvillea spectabilis branch chosen is inserted in the bottle with water, is placed in illumination room, 25 ~ 30 DEG C of temperature is maintained, 4000 ~ 5000Lux of intensity of illumination, light application time for 24 hours, irradiate 7d, choose 4 ~ 6cm Bougainvillea spectabilis sprout segments;
(3) it is surface-treated:The Bougainvillea spectabilis sprout segment gathered is rinsed well using flowing water, 10 are cleaned with 2% Tween-80 ~ 15min is rinsed well using flowing water;
(4) it sterilizes:It is impregnated using 75% alcohol, 0.1%HgCl 5 ~ 10min of solution disinfection, aseptic water washing is clean;
(5) it cuts:The Bougainvillea spectabilis sprout segment notch disinfected is updated processing, retains 2 ~ 3cm;
(6) Initial culture:The Bougainvillea spectabilis segment handled well is seeded on culture medium I, is positioned over 25 ~ 26 DEG C of temperature, per the daylight According to 20h, intensity of illumination 2000Lux, 45 ~ 50d of culture;
(7) proliferation and subculture:Selecting step(6)In obtained aseptic seedling be inoculated on culture medium II and carry out Multiplying culture, train Foster condition is:25 ~ 26 DEG C of temperature, daily illumination 12h, intensity of illumination are 5000 ~ 7000Lux, cultivate 25 ~ 30d;Repeat above-mentioned increasing Grow process;
(8) it takes root:Choose growing way stalwartness Bougainvillea spectabilis aseptic seedling, be seeded in culture medium III, be positioned over 25 ~ 26 DEG C of temperature, In daily illumination 14h, the environment that intensity of illumination is 5000 ~ 7000Lux, 25 ~ 30d is cultivated;
(9) hardening is tamed:The Bougainvillea spectabilis seedling for the well developed root system that step (8) obtains is transferred in small temperature canopy and is tamed, is shaded, It is 20 ~ 30 DEG C to keep temperature in small temperature canopy, and humid control humidity after 85 ~ 95%, 7d is decremented to 80%, hereafter gradually successively decreases, 15d Bougainvillea spectabilis seedling is removed from tissue culture medium (TCM) afterwards, the culture medium of root is cleaned up, in plantation to substrate soil.
3. a kind of Bougainvillea spectabilis detoxification method for tissue culture according to claim 1, it is characterised in that:Culture medium I changes for MS 1.0mg/LIBA, 2.0mg/LZT, 30g/L sucrose and 3.5g/L agar are added in good culture medium.
4. a kind of Bougainvillea spectabilis detoxification method for tissue culture according to claim 1, it is characterised in that:Culture medium II changes for MS 3mg/LZT, 0.1mg/LIAA, 0.5mg/LNAA, 30g/L sucrose and 3.5g/L agar are added in good culture medium.
5. a kind of Bougainvillea spectabilis detoxification method for tissue culture according to claim 1, it is characterised in that:Culture medium III changes for MS 0.5mg/LNAA, 0.2mg/LIBA, 20g/L sucrose and 3.5g/L agar are added in good culture medium.
6. a kind of Bougainvillea spectabilis detoxification method for tissue culture according to claim 1, it is characterised in that:The culture medium I, training The pH for supporting base II and culture medium II is adjusted to 5.8.
7. a kind of Bougainvillea spectabilis detoxification method for tissue culture according to claim 1, it is characterised in that:In the step (4) The alcohol disinfecting time is no more than 1min.
8. a kind of Bougainvillea spectabilis detoxification method for tissue culture according to claim 1, it is characterised in that:The step(9)In Artificially feed is carried out under the conditions of the domestication hardening link cloudy day, light filling intensity is 3000 ~ 5000Lux.
9. a kind of Bougainvillea spectabilis detoxification method for tissue culture according to claim 1, it is characterised in that:The step(9)In Hardening step is tamed, being transferred to small temperature canopy, dusk or early morning divulge information half an hour daily after 10 ~ 15 days.
CN201810661263.5A 2018-06-25 2018-06-25 A kind of Bougainvillea spectabilis detoxification method for tissue culture Pending CN108522289A (en)

Priority Applications (1)

Application Number Priority Date Filing Date Title
CN201810661263.5A CN108522289A (en) 2018-06-25 2018-06-25 A kind of Bougainvillea spectabilis detoxification method for tissue culture

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
CN201810661263.5A CN108522289A (en) 2018-06-25 2018-06-25 A kind of Bougainvillea spectabilis detoxification method for tissue culture

Publications (1)

Publication Number Publication Date
CN108522289A true CN108522289A (en) 2018-09-14

Family

ID=63487212

Family Applications (1)

Application Number Title Priority Date Filing Date
CN201810661263.5A Pending CN108522289A (en) 2018-06-25 2018-06-25 A kind of Bougainvillea spectabilis detoxification method for tissue culture

Country Status (1)

Country Link
CN (1) CN108522289A (en)

Cited By (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN109258467A (en) * 2018-10-16 2019-01-25 广西壮族自治区林业科学研究院 A method of preventing bougainvillea tissue-cultured seedling yellow
CN112673957A (en) * 2020-12-22 2021-04-20 中国热带农业科学院热带作物品种资源研究所 Rapid breeding method of bougainvillea spectabilis tissue culture plants
CN115428731A (en) * 2022-08-11 2022-12-06 厦门路桥百城建设投资有限公司 Method for rapidly inducing roots through tide-type in-vitro cutting of bougainvillea spectabilis and application of method

Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN106069753A (en) * 2016-06-16 2016-11-09 海南省农业科学院热带园艺研究所 A kind of Bougainvillea spectabilis aseptic seeding and method for quickly breeding

Patent Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN106069753A (en) * 2016-06-16 2016-11-09 海南省农业科学院热带园艺研究所 A kind of Bougainvillea spectabilis aseptic seeding and method for quickly breeding

Non-Patent Citations (7)

* Cited by examiner, † Cited by third party
Title
MICHELE ZACCAI, ET AL.: "Regeneration and transformation system in Mirabilis jalapa", 《SCIENTIA HORTICULTURAE》 *
叶顶英: "三角梅组培苗试管内外生根研究 ", 《北方园艺》 *
周祖富: "几种观赏花卉的组织培养和快速繁殖 ", 《广西农业生物科学》 *
李师翁: "叶子花的组织培养与快速繁殖 ", 《植物生理学通讯》 *
李师翁等: "叶子花的组织培养与微繁技术研究 ", 《西北植物学报》 *
田高飞等: "三角梅属(Bougainvillea)研究进展 ", 《北华大学学报(自然科学版)》 *
董永义等: "盆栽三角梅的组织培养 ", 《北方园艺》 *

Cited By (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN109258467A (en) * 2018-10-16 2019-01-25 广西壮族自治区林业科学研究院 A method of preventing bougainvillea tissue-cultured seedling yellow
CN112673957A (en) * 2020-12-22 2021-04-20 中国热带农业科学院热带作物品种资源研究所 Rapid breeding method of bougainvillea spectabilis tissue culture plants
CN115428731A (en) * 2022-08-11 2022-12-06 厦门路桥百城建设投资有限公司 Method for rapidly inducing roots through tide-type in-vitro cutting of bougainvillea spectabilis and application of method
CN115428731B (en) * 2022-08-11 2024-04-19 厦门路桥百城建设投资有限公司 Triangle Mei Chaoxi type in-vitro cutting rapid root induction method and application thereof

Similar Documents

Publication Publication Date Title
KR101587707B1 (en) Producing method of orchid seedlings
CN105475130A (en) Castanopsis hystrix high efficiency isolated culture plant regeneration method
JP6316545B2 (en) Cutting seedling production method
CN103348920B (en) Rapid propagation method for high quality seedlings of Kyara
CN106258979B (en) A kind of hybridization paulownia seedling cultural method
CN101946703B (en) Method for regenerating plants of Chinese rose by using leaves as explants
CN104322375A (en) Method for rapidly propagating dendrobium chrysotoxumLindl. seeds by tissue culture
CN103688855B (en) A kind of leaflet red bean isolated seed embryo and plant regeneration method
CN102845313A (en) Method for quickly in-vitro actinidia kolomikta propagating
CN108522289A (en) A kind of Bougainvillea spectabilis detoxification method for tissue culture
CN101133704A (en) Sterilized spore germination of platycerium wallichii and test tube seedling cultivating method
CN101502239B (en) Method for rapid propagation and cultivation of carnation seedling by tissue culture
JPH05268843A (en) Cultured species of moss and method for cultivating mosses using the same
CN103583360A (en) Method for improving Abelia seedling salt tolerance by oriented induction
CN103392602A (en) Sterile seeding and test-tube plantlet propagation method of sword-leaved cymbidiums
CN104604735A (en) Tissue culture and rapid propagation method for American lagerstroemia indica pink velour
CN104488721B (en) A kind of quick breeding method for tissue culture of snowflake grass
CN100391333C (en) Aseptic seedling tissue culturing and test tube seedling hardening off and transplating technology for anthurium andraeanum
CN106605596B (en) A method of mass propagation Lycoris aurea is occurred by body embryo
CN107466850B (en) Blueberry planting and seedling rapid cultivation method
CN112616675B (en) Tissue culture and rapid propagation method for Zingiber dance
CN104396756A (en) Culture and rapid propagation method for rhynchostylis gigantea shoot apices
CN108391591A (en) A kind of Golden Bell Tree tissue cultivation rapid breeding method
CN102763598B (en) Method for breeding wenshan paphiopedilum seedlings by using somatic embryo
CN108812327B (en) Culture solution and culture method for water culture propagation of chrysosporium

Legal Events

Date Code Title Description
PB01 Publication
PB01 Publication
SE01 Entry into force of request for substantive examination
SE01 Entry into force of request for substantive examination
WD01 Invention patent application deemed withdrawn after publication

Application publication date: 20180914

WD01 Invention patent application deemed withdrawn after publication