CN114586685A - Culture method of Chinese starjasmine stem virus-free seedlings - Google Patents
Culture method of Chinese starjasmine stem virus-free seedlings Download PDFInfo
- Publication number
- CN114586685A CN114586685A CN202210281740.1A CN202210281740A CN114586685A CN 114586685 A CN114586685 A CN 114586685A CN 202210281740 A CN202210281740 A CN 202210281740A CN 114586685 A CN114586685 A CN 114586685A
- Authority
- CN
- China
- Prior art keywords
- adventitious bud
- culture
- stem
- culturing
- culture medium
- 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
Links
- 238000012136 culture method Methods 0.000 title claims abstract description 8
- 241000574434 Jasminum laurifolium Species 0.000 title claims 2
- 230000006698 induction Effects 0.000 claims abstract description 46
- 241001300674 Plukenetia volubilis Species 0.000 claims abstract description 23
- 241000196324 Embryophyta Species 0.000 claims abstract description 10
- 238000001784 detoxification Methods 0.000 claims abstract description 6
- 238000000034 method Methods 0.000 claims abstract description 5
- 239000001963 growth medium Substances 0.000 claims description 36
- 238000012258 culturing Methods 0.000 claims description 32
- 238000005286 illumination Methods 0.000 claims description 31
- 229920001817 Agar Polymers 0.000 claims description 19
- 239000008272 agar Substances 0.000 claims description 19
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 claims description 15
- 229930006000 Sucrose Natural products 0.000 claims description 15
- 238000002791 soaking Methods 0.000 claims description 15
- 239000005720 sucrose Substances 0.000 claims description 15
- 230000035755 proliferation Effects 0.000 claims description 14
- 239000011159 matrix material Substances 0.000 claims description 12
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 10
- 239000008223 sterile water Substances 0.000 claims description 10
- 238000005406 washing Methods 0.000 claims description 10
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 10
- 238000011081 inoculation Methods 0.000 claims description 9
- 230000003203 everyday effect Effects 0.000 claims description 7
- 210000003608 fece Anatomy 0.000 claims description 5
- 239000010871 livestock manure Substances 0.000 claims description 5
- 229960002523 mercuric chloride Drugs 0.000 claims description 5
- LWJROJCJINYWOX-UHFFFAOYSA-L mercury dichloride Chemical compound Cl[Hg]Cl LWJROJCJINYWOX-UHFFFAOYSA-L 0.000 claims description 5
- 239000003415 peat Substances 0.000 claims description 5
- 239000012883 rooting culture medium Substances 0.000 claims description 5
- 239000012882 rooting medium Substances 0.000 claims description 5
- 239000004576 sand Substances 0.000 claims description 5
- 239000002689 soil Substances 0.000 claims description 5
- 230000001954 sterilising effect Effects 0.000 claims description 5
- 240000005779 Jasminum multiflorum Species 0.000 abstract description 14
- 235000013399 edible fruits Nutrition 0.000 abstract description 8
- DTOSIQBPPRVQHS-PDBXOOCHSA-N alpha-linolenic acid Chemical compound CC\C=C/C\C=C/C\C=C/CCCCCCCC(O)=O DTOSIQBPPRVQHS-PDBXOOCHSA-N 0.000 abstract description 4
- WRIDQFICGBMAFQ-UHFFFAOYSA-N (E)-8-Octadecenoic acid Natural products CCCCCCCCCC=CCCCCCCC(O)=O WRIDQFICGBMAFQ-UHFFFAOYSA-N 0.000 abstract description 2
- LQJBNNIYVWPHFW-UHFFFAOYSA-N 20:1omega9c fatty acid Natural products CCCCCCCCCCC=CCCCCCCCC(O)=O LQJBNNIYVWPHFW-UHFFFAOYSA-N 0.000 abstract description 2
- QSBYPNXLFMSGKH-UHFFFAOYSA-N 9-Heptadecensaeure Natural products CCCCCCCC=CCCCCCCCC(O)=O QSBYPNXLFMSGKH-UHFFFAOYSA-N 0.000 abstract description 2
- 244000105624 Arachis hypogaea Species 0.000 abstract description 2
- 235000010777 Arachis hypogaea Nutrition 0.000 abstract description 2
- OYHQOLUKZRVURQ-HZJYTTRNSA-N Linoleic acid Chemical compound CCCCC\C=C/C\C=C/CCCCCCCC(O)=O OYHQOLUKZRVURQ-HZJYTTRNSA-N 0.000 abstract description 2
- 239000005642 Oleic acid Substances 0.000 abstract description 2
- ZQPPMHVWECSIRJ-UHFFFAOYSA-N Oleic acid Natural products CCCCCCCCC=CCCCCCCCC(O)=O ZQPPMHVWECSIRJ-UHFFFAOYSA-N 0.000 abstract description 2
- 241000700605 Viruses Species 0.000 abstract description 2
- 235000020661 alpha-linolenic acid Nutrition 0.000 abstract description 2
- 239000002537 cosmetic Substances 0.000 abstract description 2
- 229940079593 drug Drugs 0.000 abstract description 2
- 239000003814 drug Substances 0.000 abstract description 2
- 239000004519 grease Substances 0.000 abstract description 2
- 238000003306 harvesting Methods 0.000 abstract description 2
- QXJSBBXBKPUZAA-UHFFFAOYSA-N isooleic acid Natural products CCCCCCCC=CCCCCCCCCC(O)=O QXJSBBXBKPUZAA-UHFFFAOYSA-N 0.000 abstract description 2
- 229960004232 linoleic acid Drugs 0.000 abstract description 2
- 229960004488 linolenic acid Drugs 0.000 abstract description 2
- ZQPPMHVWECSIRJ-KTKRTIGZSA-N oleic acid Chemical compound CCCCCCCC\C=C/CCCCCCCC(O)=O ZQPPMHVWECSIRJ-KTKRTIGZSA-N 0.000 abstract description 2
- 229960002969 oleic acid Drugs 0.000 abstract description 2
- 235000021085 polyunsaturated fats Nutrition 0.000 abstract description 2
- 238000002054 transplantation Methods 0.000 abstract description 2
- 235000003276 Apios tuberosa Nutrition 0.000 abstract 1
- 235000010744 Arachis villosulicarpa Nutrition 0.000 abstract 1
- 235000015489 Emblica officinalis Nutrition 0.000 abstract 1
- 240000000908 Phyllanthus acidus Species 0.000 abstract 1
- 230000004083 survival effect Effects 0.000 description 3
- 235000017060 Arachis glabrata Nutrition 0.000 description 1
- 235000018262 Arachis monticola Nutrition 0.000 description 1
- 240000007581 Derris trifoliata Species 0.000 description 1
- 241001026836 Docynia indica Species 0.000 description 1
- 244000288157 Passiflora edulis Species 0.000 description 1
- 235000000370 Passiflora edulis Nutrition 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 235000020232 peanut Nutrition 0.000 description 1
- 238000004161 plant tissue culture Methods 0.000 description 1
Classifications
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
- A01H4/00—Plant reproduction by tissue culture techniques ; Tissue culture techniques therefor
- A01H4/008—Methods for regeneration to complete plants
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
- A01H4/00—Plant reproduction by tissue culture techniques ; Tissue culture techniques therefor
- A01H4/002—Culture media for tissue culture
-
- 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
Landscapes
- Life Sciences & Earth Sciences (AREA)
- Developmental Biology & Embryology (AREA)
- Engineering & Computer Science (AREA)
- Biotechnology (AREA)
- Cell Biology (AREA)
- Botany (AREA)
- Environmental Sciences (AREA)
- Breeding Of Plants And Reproduction By Means Of Culturing (AREA)
Abstract
The invention discloses a culture method of a Chinese starjasmine stem virus-free seedling, wherein the Chinese starjasmine stem is also called as plukenetia volubilis, Indian gooseberry, Indian groundnut and the like, and the grease squeezed from the Chinese starjasmine stem seeds is rich in polyunsaturated fat containing alpha-linolenic acid, alpha-linoleic acid, oleic acid and the like, and belongs to the fields of medicines, cosmetics and the like. The plukenetia volubilis linneo is a perennial vine plant, and can continuously harvest fruits for many years after being planted once, but since plukenetia volubilis linneo seedlings carry viruses, the later-stage plants are seriously damaged, so that the fruits fall and the fruit yield is reduced. Therefore, the invention uses the stem tip of the plukenetia volubilis linneo plant as an explant, obtains a large batch of plukenetia volubilis linneo detoxified seedlings through the processes of adventitious bud induction, detoxification treatment, propagation culture, rooting culture, hardening seedling transplantation and the like, effectively reduces the morbidity in the later period of field planting, and improves the fruit yield of the plukenetia volubilis linneo.
Description
Technical Field
The invention relates to the field of plant tissue culture, and particularly discloses a culture method of a detoxified seedling of plukenetia volubilis linneo.
Background
The Plukenetia volubilis L is also called as Plukenetia volubilis, Indian crabapple, Indian peanut and the like, and the grease squeezed from the seeds of the Plukenetia volubilis is rich in polyunsaturated fat containing alpha-linolenic acid, alpha-linoleic acid, oleic acid and the like, and is used in the fields of medicines, cosmetics and the like. The plukenetia volubilis linneo is introduced into the west double banna tropical plantations of Chinese academy of sciences in 2006 to be cultivated and planted successfully, and the plukenetia volubilis linneo industry is vigorously developed in provinces and cities of Yunnan, Guizhou and the like at present. The plukenetia volubilis linneo is a perennial vine plant, and can continuously harvest fruits for many years after being planted once, but since plukenetia volubilis linneo seedlings carry viruses, the later-stage plants are seriously damaged, so that the fruits fall and the fruit yield is reduced. Therefore, establishing an effective passion fruit virus-free seedling culture method has important significance for the development of the plukenetia volubilis linneo industry.
Disclosure of Invention
The invention aims to provide a culture method of a Chinese starjasmine virus-free seedling, which takes the stem tip of a Chinese starjasmine plant as an explant, and obtains a large amount of Chinese starjasmine virus-free seedlings through the processes of adventitious bud induction, virus-free treatment, propagation culture, rooting culture, hardening seedling transplantation and the like, thereby effectively reducing the morbidity of the Chinese starjasmine in the later period of field planting, improving the fruit yield of the Chinese starjasmine and achieving the aim of the invention.
The invention relates to a culture method of a derris trifoliata virus-free seedling, which comprises the following process steps:
(1) adventitious bud induction: selecting 6-8 cm stem tips of plukenetia volubilis linneo plants as explants, sterilizing the explants for 10s by using 75% ethanol, washing the explants for 5 times by using sterile water, soaking the washed explants in 0.1% mercuric chloride solution for 6min, washing the washed explants for 6 times by using sterile water, cutting the washed shoots into 2-3 cm stem segments with nodes, inoculating the stem segments into adventitious bud induction culture, and carrying out dark culture at 25 ℃ until adventitious buds are formed, wherein the adventitious bud induction culture medium is MS + 0.1-0.5 mg/L TDZ + 1.0-2.0 mg/L6-BA + 0.1-0.5 mg/L NAA + 15-30 g/L sucrose + 3.5-6.0 g/L agar, and the pH value is 5.4-5.8;
(2) detoxification treatment: under the aseptic condition, soaking aseptic filter paper in a sterilized agar-free adventitious bud induction culture medium for 10-20 min, flatly paving the aseptic filter paper in an aseptic culture bottle, tightly attaching the adventitious bud obtained in the step (1) to the filter paper, culturing in the dark at 40-45 ℃ for 3-5 d, stripping the stem tip of a new adventitious bud by 0.5cm, inoculating the stem tip into the adventitious bud induction culture medium, and culturing for 20-30 d at 25 ℃ for 12-15 h each day under the illumination intensity of 1500-2000 lx to form the new adventitious bud. And then soaking sterile filter paper in a sterilized agar-free adventitious bud induction culture medium for 10-20 min, flatly paving the sterile filter paper in a sterile culture bottle, tightly attaching the newly formed adventitious bud to the filter paper, culturing for 3-5 d at 40-45 ℃ in the dark, stripping the stem tip of 0.2cm of the newly formed adventitious bud, inoculating the stem tip into the adventitious bud induction culture medium, and culturing for 20-30 d at 25 ℃ under the conditions of illumination intensity of 1500-2000 lx and illumination intensity of 12-15 h every day to form a new adventitious bud.
(3) And (3) proliferation culture: cutting the adventitious bud obtained in the step (2) from the base part, inoculating the cut adventitious bud to a multiplication culture medium for multiplication culture, and culturing under the conditions of illumination intensity of 1500-2000 lx and culture temperature of 25 ℃ for 12-15 h every day after inoculation, wherein the transfer is performed every 20-30 d. The enrichment culture medium is MS + 1.0-2.0 mg/L6-BA + 0.1-0.5 mg/L NAA + 15-30 g/L sucrose + 3.5-6.0 g/L agar, and the pH value is 5.4-5.8;
(4) rooting culture: and (4) cutting the adventitious bud which is obtained in the step (3) and grows to 2-4 cm, inoculating the cut adventitious bud to a rooting culture medium for rooting culture, after inoculation, culturing the bud in the dark at 25 ℃ for 3-5 days, and then culturing the bud under the conditions of illumination intensity of 2000-3000 lx and culture temperature of 25 ℃ for rooting every day for 12-15 hours. The rooting medium is 1/2MS + 0.1-0.5 mg/L IBA + 1.0-2.0 mg/L GGR + 15-30 g/L sucrose + 3.5-6.0 g/L agar, and the pH value is 5.4-5.8;
(5) hardening and transplanting seedlings: and (5) cultivating the rooted seedlings obtained in the step (4) in a mixed matrix to form seedlings, and obtaining the detoxified seedlings of the plukenetia volubilis linneo. The mixed matrix is formed by mixing peat soil, farmyard manure and river sand according to the ratio of 8:1: 1.
The method has the characteristics of low cost, good effect, strong operability and the like, can be used for industrially producing the Chinese starjasmine stem virus-free seedlings in large batch, and can effectively reduce the disease rate of the Chinese starjasmine stem after the virus-free seedlings are planted, improve the fruiting rate and improve the economic benefit of the Chinese starjasmine stem.
Detailed Description
The following examples are further illustrative of the present invention and are not intended to be limiting thereof.
Example one
(1) Adventitious bud induction: selecting 6-8 cm stem tips of plukenetia volubilis linneo plants as explants, sterilizing the explants for 10s by using 75% ethanol, washing the explants for 5 times by using sterile water, soaking the washed explants in 0.1% mercuric chloride solution for 6min, washing the washed explants for 6 times by using the sterile water, cutting the washed explants by using sterile filter paper into 2-3 cm stem segments with nodes, inoculating the stem segments with the nodes into adventitious bud induction culture, carrying out dark culture at 25 ℃ for 20 days to form adventitious buds, wherein the pollution rate is lower than 5%, and the adventitious bud induction rate can reach 94.8%. The adventitious bud induction culture medium is MS +0.1mg/L TDZ +1.0 mg/L6-BA +0.1mg/L NAA +15g/L sucrose +3.5g/L agar, and the pH value is 5.4;
(2) detoxification treatment: under the aseptic condition, soaking sterile filter paper in a sterilized agar-free adventitious bud induction culture medium for 10min, flatly paving the sterile filter paper in a sterile culture bottle, tightly attaching the adventitious bud obtained in the step (1) to the filter paper, culturing in the dark at 40 ℃ for 3d, stripping the stem tip of a new adventitious bud of 0.5cm, inoculating the stem tip into the adventitious bud induction culture medium, and culturing for 20d at 25 ℃ for 12h per day under the illumination intensity of 1500-2000 lx to form a new adventitious bud with the adventitious bud induction rate of 75.3%. And then soaking sterile filter paper in a sterilized adventitious bud induction culture medium without agar for 10min, flatly paving the sterile filter paper in a sterile culture bottle, tightly attaching the newly formed adventitious bud to the filter paper, culturing for 3d at 40 ℃ in the dark, stripping the stem tip of 0.2cm of the newly formed adventitious bud, inoculating the stem tip into the adventitious bud induction culture medium, and culturing for 20d at 25 ℃ for 12h each day under the conditions that the illumination intensity is 1500lx and the culture temperature is 25 ℃ to form a new adventitious bud, wherein the adventitious bud induction rate is 51.8%.
(3) And (3) proliferation culture: cutting the adventitious bud obtained in the step (2) from the base part, inoculating the cut adventitious bud to an enrichment culture medium for enrichment culture, and culturing under the conditions of illumination intensity of 1500lx and culture temperature of 25 ℃ for 12h every day after inoculation, wherein the inoculation is carried out once every 20d, and the multiplication coefficient is 8.09. The proliferation culture medium is MS +1.0 mg/L6-BA +0.1mg/L NAA +15g/L sucrose +3.5g/L agar, and the pH value is 5.4;
(4) rooting culture: and (3) cutting the adventitious bud growing to 2-4 cm obtained in the step (3), inoculating the cut adventitious bud to a rooting culture medium for rooting culture, after inoculation, culturing the bud in the dark at 25 ℃ for 3 days, then placing the bud under the conditions of 12 hours of illumination each day, the illumination intensity being 2000lx and the culture temperature being 25 ℃ for 10 days, and then beginning rooting, wherein the rooting rate reaches 95.2%. The rooting medium is 1/2MS +0.1mg/L IBA +1.0mg/L GGR +15g/L sucrose +3.5g/L agar, and the pH value is 5.4;
(5) hardening and transplanting seedlings: and (5) cultivating the rooted seedlings obtained in the step (4) in a mixed matrix to form seedlings to obtain the Chinese starjasmine stem virus-free seedlings, wherein the transplanting survival rate reaches 97.6%. The mixed matrix is formed by mixing peat soil, farmyard manure and river sand according to the ratio of 8:1: 1. After the seedlings are transplanted to a field for planting, the 3 rd year morbidity is lower than 2.3%.
Example two
(1) Adventitious bud induction: selecting 6-8 cm stem tips of plukenetia volubilis linneo plants as explants, sterilizing the explants for 10s by using 75% ethanol, washing the explants for 5 times by using sterile water, soaking the washed explants in 0.1% mercuric chloride solution for 6min, washing the washed explants for 6 times by using the sterile water, cutting the washed explants by using sterile filter paper into 2-3 cm stem segments with nodes, inoculating the stem segments with the nodes into adventitious bud induction culture, carrying out dark culture at 25 ℃ for 18 days to form adventitious buds, wherein the pollution rate is lower than 4%, and the adventitious bud induction rate can reach 96.1%. The adventitious bud induction culture medium is MS +0.mg/L TDZ +1.5 mg/L6-BA +0.3mg/L NAA +17g/L sucrose +4.5g/L agar, and the pH value is 5.6;
(2) detoxification treatment: under the aseptic condition, soaking sterile filter paper in a sterilized agar-free adventitious bud induction culture medium for 15min, flatly paving the sterile filter paper in a sterile culture bottle, tightly attaching the adventitious bud obtained in the step (1) to the filter paper, culturing for 4d at 43 ℃ in the dark, stripping the stem tip of a new adventitious bud by 0.5cm, inoculating the stem tip into the adventitious bud induction culture medium, and culturing for 25d at 25 ℃ under the conditions of 14h of illumination each day, the illumination intensity of 1500-2000 lx and the culture temperature of 25 ℃ to form a new adventitious bud, wherein the adventitious bud induction rate is 73.8%. And then soaking sterile filter paper in a sterilized adventitious bud induction culture medium without agar for 15min, flatly paving the sterile filter paper in a sterile culture bottle, tightly attaching the newly formed adventitious bud to the filter paper, culturing for 4d at 43 ℃ in the dark, stripping the stem tip of 0.2cm of the newly formed adventitious bud, inoculating the stem tip into the adventitious bud induction culture medium, and culturing for 25d at 25 ℃ under the conditions of 14h of illumination each day, the illumination intensity of 1700lx and the culture temperature of 25 ℃ to form a new adventitious bud, wherein the adventitious bud induction rate is 54.6%. .
(3) And (3) proliferation culture: cutting the adventitious bud obtained in the step (2) from the base part, inoculating the cut adventitious bud to a proliferation culture medium for proliferation culture, and culturing under the conditions of illumination intensity of 1700lx and culture temperature of 25 ℃ for 13h every day, wherein the proliferation coefficient is 8.27 after the culture is carried out every 25 d. The proliferation culture medium is MS +1.5 mg/L6-BA +0.3mg/L NAA +17g/L sucrose +4.5g/L agar, and the pH value is 5.6;
(4) rooting culture: and (3) cutting the adventitious bud growing to 2-4 cm obtained in the step (3), inoculating the cut adventitious bud to a rooting culture medium for rooting culture, culturing in the dark at 25 ℃ for 3-5 days after inoculation, then placing the culture medium under the condition of illumination for 14h each day, wherein the illumination intensity is 2500lx, the culture temperature is 25 ℃ and the culture time is 10 days, and the rooting rate reaches 100%. The rooting medium is 1/2MS +0.3mg/L IBA +1.5mg/L GGR +18g/L sucrose +4.0g/L agar, and the pH value is 5.6;
(5) hardening and transplanting seedlings: and (5) cultivating the rooted seedlings obtained in the step (4) into seedlings in a mixed matrix to obtain the detoxified seedlings of the plukenetia volubilis linneo, wherein the transplanting survival rate reaches 99.5%, and the mixed matrix is formed by mixing peat soil, farmyard manure and river sand according to the ratio of 8:1: 1. After the seedlings are transplanted to a field for planting, the 3 rd year morbidity is lower than 2.5%.
Example four
(1) Adventitious bud induction: selecting 6-8 cm stem tips of a plukenetia volubilis linneo plant as explants, sterilizing the explants by 75% ethanol for 10s, washing the explants for 5 times by using sterile water, soaking the washed explants in 0.1% mercuric chloride solution for 6min, washing the washed explants by using the sterile water for 6 times, cutting the washed explants by using sterile filter paper into 2-3 cm stem segments with nodes, inoculating the stem segments with the segments into adventitious bud induction culture, and performing dark culture at 25 ℃ for 19 days to form adventitious buds, wherein the pollution rate is less than 3%, the adventitious bud induction rate can reach 97.8%, the adventitious bud induction culture medium is MS +0.5mg/L TDZ +2.0 mg/L6-BA +0.5mg/L NAA +30g/L sucrose +6.0g/L agar, and the pH is 5.8;
(2) detoxification treatment: under the aseptic condition, soaking aseptic filter paper in a sterilized adventitious bud induction culture medium without agar for 20min, flatly paving the aseptic filter paper in an aseptic culture bottle, tightly attaching the adventitious bud obtained in the step (1) to the filter paper, culturing in the dark at 45 ℃ for 3-5 d, stripping the stem tip of a new adventitious bud by 0.5cm, inoculating the stem tip into the adventitious bud induction culture medium, and culturing for 30d at 25 ℃ under the conditions of 15h of illumination each day, the illumination intensity of 2000lx and the culture temperature to form a new adventitious bud, wherein the adventitious bud induction rate is 80.4%. . And then soaking sterile filter paper in a sterilized adventitious bud induction culture medium without agar for 20min, flatly paving the sterile filter paper in a sterile culture bottle, tightly attaching the newly formed adventitious bud to the filter paper, culturing for 5d at 45 ℃ in the dark, stripping the stem tip of the newly formed adventitious bud by 0.2cm, inoculating the stem tip into the adventitious bud induction culture medium, and culturing for 30d at 25 ℃ under the conditions of 15h of illumination each day, the illumination intensity of 2000lx and the culture temperature of 62 ℃ to form a new adventitious bud with the adventitious bud induction rate of 62.8%. .
(3) And (3) proliferation culture: cutting the adventitious bud obtained in the step (2) from the base part, inoculating the cut adventitious bud to a proliferation culture medium for proliferation culture, culturing the cut adventitious bud under the conditions of illumination intensity of 1500-2000 lx and culture temperature of 25 ℃ for 15 hours every day after inoculation, and transferring the cut adventitious bud once every 30 days until the proliferation coefficient reaches 7.8. The proliferation culture medium is MS +2.0 mg/L6-BA +0.5mg/L NAA +30g/L sucrose +6.0g/L agar, and the pH value is 5.8;
(4) rooting culture: and (3) cutting the adventitious bud growing to 2-4 cm obtained in the step (3), inoculating the cut adventitious bud to a rooting culture medium for rooting culture, after inoculation, culturing the bud in the dark at 25 ℃ for 5 days, then placing the bud under the conditions of 15 hours of illumination each day, the illumination intensity being 3000lx and the culture temperature being 25 ℃ for 9 days, and then beginning rooting, wherein the rooting rate reaches 100%. The rooting medium is 1/2MS +0.5mg/L IBA +2.0mg/L GGR +30g/L sucrose +6.0g/L agar, and the pH value is 5.8;
(5) hardening and transplanting seedlings: and (5) cultivating the rooted seedlings obtained in the step (4) in a mixed matrix to form seedlings to obtain the Chinese starjasmine stem virus-free seedlings, wherein the transplanting survival rate reaches 98%. The mixed matrix is formed by mixing peat soil, farmyard manure and river sand according to the ratio of 8:1:1, and the 3 rd year morbidity of the mixed matrix after the mixed matrix is transplanted to a field for planting is lower than 2.0%.
Claims (1)
1. A culture method of a Chinese starjasmine stem virus-free seedling is characterized by comprising the following process steps:
(1) adventitious bud induction: selecting 6-8 cm stem tips of plukenetia volubilis linneo plants as explants, sterilizing the explants for 10s by using 75% ethanol, washing the explants for 5 times by using sterile water, soaking the washed explants in 0.1% mercuric chloride solution for 6min, washing the washed explants for 6 times by using sterile water, cutting the washed shoots into 2-3 cm stem segments with nodes, inoculating the stem segments into adventitious bud induction culture, and carrying out dark culture at 25 ℃ until adventitious buds are formed, wherein the adventitious bud induction culture medium is MS + 0.1-0.5 mg/L TDZ + 1.0-2.0 mg/L6-BA + 0.1-0.5 mg/L NAA + 15-30 g/L sucrose + 3.5-6.0 g/L agar, and the pH value is 5.4-5.8;
(2) detoxification treatment: under the aseptic condition, soaking aseptic filter paper in a sterilized agar-free adventitious bud induction culture medium for 10-20 min, flatly paving the aseptic filter paper in an aseptic culture bottle, tightly attaching the adventitious bud obtained in the step (1) to the filter paper, culturing in the dark at 40-45 ℃ for 3-5 d, stripping the stem tip of a new adventitious bud by 0.5cm, inoculating the stem tip into the adventitious bud induction culture medium, and culturing for 20-30 d at 25 ℃ for 12-15 h each day under the illumination intensity of 1500-2000 lx to form a new adventitious bud; soaking sterile filter paper in a sterilized agar-free adventitious bud induction culture medium for 10-20 min, flatly paving the sterile filter paper in a sterile culture bottle, tightly attaching the newly formed adventitious bud to the filter paper, culturing in the dark at 40-45 ℃ for 3-5 d, stripping a stem tip of 0.2cm of the newly formed adventitious bud, inoculating the stem tip into the adventitious bud induction culture medium, and culturing at 25 ℃ for 20-30 d to form a new adventitious bud under the conditions of illumination for 12-15 h every day, illumination intensity of 1500-2000 lx and culture temperature of 25 ℃;
(3) and (3) proliferation culture: cutting the adventitious bud obtained in the step (2) from the base part and inoculating the adventitious bud onto an enrichment culture medium for enrichment culture, and culturing under the conditions of illumination for 12-15 hours each day, illumination intensity of 1500-2000 lx and culture temperature of 25 ℃, wherein the adventitious bud is transferred once every 20-30 days; the enrichment culture medium is MS + 1.0-2.0 mg/L6-BA + 0.1-0.5 mg/L NAA + 15-30 g/L sucrose + 3.5-6.0 g/L agar, and the pH value is 5.4-5.8;
(4) rooting culture: cutting the adventitious bud which is obtained in the step (3) and grows to 2-4 cm in length, inoculating the cut adventitious bud to a rooting culture medium for rooting culture, after inoculation, culturing the bud in the dark at 25 ℃ for 3-5 days, and then culturing the bud under the conditions of illumination for 12-15 hours each day, the illumination intensity being 2000-3000 lx and the culture temperature being 25 ℃ until the bud grows to root; the rooting medium is 1/2MS + 0.1-0.5 mg/LIBA + 1.0-2.0 mg/L GGR + 15-30 g/L sucrose + 3.5-6.0 g/L agar, and the pH value is 5.4-5.8;
(5) hardening and transplanting seedlings: cultivating the rooted seedlings obtained in the step (4) into seedlings in a mixed matrix to obtain the detoxified seedlings of the plukenetia volubilis linneo; the mixed matrix is formed by mixing peat soil, farmyard manure and river sand according to the ratio of 8:1: 1.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202210281740.1A CN114586685A (en) | 2022-03-21 | 2022-03-21 | Culture method of Chinese starjasmine stem virus-free seedlings |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202210281740.1A CN114586685A (en) | 2022-03-21 | 2022-03-21 | Culture method of Chinese starjasmine stem virus-free seedlings |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN114586685A true CN114586685A (en) | 2022-06-07 |
Family
ID=81809893
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202210281740.1A Pending CN114586685A (en) | 2022-03-21 | 2022-03-21 | Culture method of Chinese starjasmine stem virus-free seedlings |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN114586685A (en) |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101138319A (en) * | 2007-09-28 | 2008-03-12 | 中国科学院华南植物园 | Tristellateia australasiae tissue culturing and propagating method |
| CN104798684A (en) * | 2015-04-25 | 2015-07-29 | 玉林师范学院 | Tissue culture rapid propagation method for plukenetia volubilis L. |
| CN106342689A (en) * | 2016-09-06 | 2017-01-25 | 海南大学 | Method for quickly propagating plukenetia volubilis linneo |
-
2022
- 2022-03-21 CN CN202210281740.1A patent/CN114586685A/en active Pending
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101138319A (en) * | 2007-09-28 | 2008-03-12 | 中国科学院华南植物园 | Tristellateia australasiae tissue culturing and propagating method |
| CN104798684A (en) * | 2015-04-25 | 2015-07-29 | 玉林师范学院 | Tissue culture rapid propagation method for plukenetia volubilis L. |
| CN106342689A (en) * | 2016-09-06 | 2017-01-25 | 海南大学 | Method for quickly propagating plukenetia volubilis linneo |
Non-Patent Citations (1)
| Title |
|---|
| 唐世裔等编著: "《特色水果高产优质栽培新技术》", 28 February 2017, 湖南科学技术出版社 * |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN104067939A (en) | Tissue culture rapid propagation method of radix gentianae | |
| CN105123529A (en) | Rapid propagation and efficient cultivation method of Bletilla striata | |
| CN111280056A (en) | Subculture breeding method of stingless pepper tissue culture seedlings | |
| CN103430845A (en) | Strawberry tissue culturing method | |
| CN112273231A (en) | Method for inducing proliferation of asparagus cochinchinensis cluster buds and plant regeneration | |
| CN101926284B (en) | Monkshood-tuber tissue culture and rapid propagation method | |
| CN105191792A (en) | Intermediate propagation method of vernonia amygdalina | |
| CN107896990B (en) | Sterile germination and rapid propagation method for hibiscus syriacus seeds in dry land | |
| CN112425506A (en) | Tissue culture rapid propagation method of artemisia anomala | |
| CN104686344A (en) | Tissue culture method of liriope muscari | |
| CN103548695A (en) | Tissue culture and rapid propagation method for corydalis saxicola bunting | |
| CN110679489A (en) | Rapid propagation method of traditional Chinese medicine plant ancient uncaria | |
| CN114586685A (en) | Culture method of Chinese starjasmine stem virus-free seedlings | |
| CN114982633A (en) | Rapid propagation method of national medicine thymifoious euphorbia herb | |
| CN109863998B (en) | Tissue culture method of tilia mongolica seedlings | |
| CN107889745B (en) | Tissue culture method for preventing fir test-tube plantlet from deviating from crown | |
| CN105191808A (en) | Tissue culture and rapid propagation method for aspidistra | |
| CN111280058A (en) | Rapid breeding method of detoxified seedlings of stem tips of salvia miltiorrhiza bunge | |
| CN110679486A (en) | Culture method of curcuma zedoary tissue culture seedlings | |
| CN113068614B (en) | Method for rapidly propagating high-quality seedlings of golden strelitzia | |
| CN111248087B (en) | Tissue culture rapid propagation method for primary seedling formation of radix glehniae | |
| CN111264393B (en) | Method for rapidly breeding epimedium test-tube plantlets | |
| CN104770299B (en) | Method for sterilely germinating gentianella acuta seeds | |
| CN115735771B (en) | Efficient in-vitro propagation method for radix stemonae | |
| CN117502246B (en) | Construction method of lotus rapid propagation system |
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 | ||
| RJ01 | Rejection of invention patent application after publication |
Application publication date: 20220607 |
|
| RJ01 | Rejection of invention patent application after publication |