CN116458428B - Tissue culture seedling method for momordica grosvenori stem segments - Google Patents
Tissue culture seedling method for momordica grosvenori stem segments Download PDFInfo
- Publication number
- CN116458428B CN116458428B CN202310426135.3A CN202310426135A CN116458428B CN 116458428 B CN116458428 B CN 116458428B CN 202310426135 A CN202310426135 A CN 202310426135A CN 116458428 B CN116458428 B CN 116458428B
- Authority
- CN
- China
- Prior art keywords
- momordica grosvenori
- tissue culture
- buds
- seedlings
- bud
- 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.)
- Active
Links
- 235000011171 Thladiantha grosvenorii Nutrition 0.000 title claims abstract description 47
- 238000000034 method Methods 0.000 title claims abstract description 20
- 241001409321 Siraitia grosvenorii Species 0.000 title abstract 7
- 230000006698 induction Effects 0.000 claims abstract description 24
- 239000001963 growth medium Substances 0.000 claims abstract description 17
- 230000035755 proliferation Effects 0.000 claims abstract description 16
- 238000012258 culturing Methods 0.000 claims abstract description 15
- 239000011159 matrix material Substances 0.000 claims abstract description 10
- 238000005520 cutting process Methods 0.000 claims abstract description 7
- 239000012883 rooting culture medium Substances 0.000 claims abstract description 6
- 230000001954 sterilising effect Effects 0.000 claims abstract description 6
- 230000001939 inductive effect Effects 0.000 claims abstract description 3
- 244000185386 Thladiantha grosvenorii Species 0.000 claims description 40
- 229920001817 Agar Polymers 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 19
- 229930006000 Sucrose Natural products 0.000 claims description 19
- 239000008272 agar Substances 0.000 claims description 19
- 239000005720 sucrose Substances 0.000 claims description 19
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 19
- PRPINYUDVPFIRX-UHFFFAOYSA-N 1-naphthaleneacetic acid Chemical compound C1=CC=C2C(CC(=O)O)=CC=CC2=C1 PRPINYUDVPFIRX-UHFFFAOYSA-N 0.000 claims description 16
- 239000002609 medium Substances 0.000 claims description 16
- 239000000203 mixture Substances 0.000 claims description 14
- 239000000706 filtrate Substances 0.000 claims description 12
- 238000005286 illumination Methods 0.000 claims description 11
- 235000009814 Luffa aegyptiaca Nutrition 0.000 claims description 10
- 241000219138 Luffa Species 0.000 claims description 7
- 235000003956 Luffa Nutrition 0.000 claims description 7
- 238000009835 boiling Methods 0.000 claims description 7
- 235000019362 perlite Nutrition 0.000 claims description 7
- 239000010451 perlite Substances 0.000 claims description 7
- 235000019354 vermiculite Nutrition 0.000 claims description 7
- 239000010455 vermiculite Substances 0.000 claims description 7
- 229910052902 vermiculite Inorganic materials 0.000 claims description 7
- 238000001914 filtration Methods 0.000 claims description 6
- JTEDVYBZBROSJT-UHFFFAOYSA-N indole-3-butyric acid Chemical compound C1=CC=C2C(CCCC(=O)O)=CNC2=C1 JTEDVYBZBROSJT-UHFFFAOYSA-N 0.000 claims description 6
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 claims description 5
- 230000003203 everyday effect Effects 0.000 claims description 5
- 238000002156 mixing Methods 0.000 claims description 4
- 238000002360 preparation method Methods 0.000 claims description 4
- 239000007640 basal medium Substances 0.000 claims description 3
- 244000280244 Luffa acutangula Species 0.000 claims 1
- 238000004519 manufacturing process Methods 0.000 abstract description 4
- 238000000338 in vitro Methods 0.000 abstract description 3
- 230000008929 regeneration Effects 0.000 abstract description 3
- 238000011069 regeneration method Methods 0.000 abstract description 3
- 244000302544 Luffa aegyptiaca Species 0.000 description 9
- 238000005406 washing Methods 0.000 description 8
- NWBJYWHLCVSVIJ-UHFFFAOYSA-N N-benzyladenine Chemical compound N=1C=NC=2NC=NC=2C=1NCC1=CC=CC=C1 NWBJYWHLCVSVIJ-UHFFFAOYSA-N 0.000 description 7
- 239000008223 sterile water Substances 0.000 description 7
- 239000000463 material Substances 0.000 description 5
- 230000004083 survival effect Effects 0.000 description 4
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 3
- 235000013399 edible fruits Nutrition 0.000 description 3
- 239000012882 rooting medium Substances 0.000 description 3
- 241000219104 Cucurbitaceae Species 0.000 description 2
- 241000700605 Viruses Species 0.000 description 2
- 230000000249 desinfective effect Effects 0.000 description 2
- 238000011161 development Methods 0.000 description 2
- 230000018109 developmental process Effects 0.000 description 2
- 201000010099 disease Diseases 0.000 description 2
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 2
- 239000003814 drug Substances 0.000 description 2
- 238000005516 engineering process Methods 0.000 description 2
- 230000002349 favourable effect Effects 0.000 description 2
- 235000013305 food Nutrition 0.000 description 2
- 229960002523 mercuric chloride Drugs 0.000 description 2
- LWJROJCJINYWOX-UHFFFAOYSA-L mercury dichloride Chemical compound Cl[Hg]Cl LWJROJCJINYWOX-UHFFFAOYSA-L 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 239000012286 potassium permanganate Substances 0.000 description 2
- 230000001737 promoting effect Effects 0.000 description 2
- 239000002994 raw material Substances 0.000 description 2
- 238000004659 sterilization and disinfection Methods 0.000 description 2
- 239000008399 tap water Substances 0.000 description 2
- 235000020679 tap water Nutrition 0.000 description 2
- 230000009286 beneficial effect Effects 0.000 description 1
- 238000004140 cleaning Methods 0.000 description 1
- 238000013461 design Methods 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
- 238000004851 dishwashing Methods 0.000 description 1
- 238000004090 dissolution Methods 0.000 description 1
- 239000012153 distilled water Substances 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 239000003337 fertilizer Substances 0.000 description 1
- 235000003599 food sweetener Nutrition 0.000 description 1
- 235000013376 functional food Nutrition 0.000 description 1
- 230000002209 hydrophobic effect Effects 0.000 description 1
- 238000009776 industrial production Methods 0.000 description 1
- 230000014759 maintenance of location Effects 0.000 description 1
- 230000003020 moisturizing effect Effects 0.000 description 1
- 239000006870 ms-medium Substances 0.000 description 1
- 235000016709 nutrition Nutrition 0.000 description 1
- 230000035764 nutrition Effects 0.000 description 1
- 229910052700 potassium Inorganic materials 0.000 description 1
- 239000000843 powder Substances 0.000 description 1
- 238000003825 pressing Methods 0.000 description 1
- 238000010008 shearing Methods 0.000 description 1
- 238000002791 soaking Methods 0.000 description 1
- 238000005728 strengthening Methods 0.000 description 1
- 239000003765 sweetening agent Substances 0.000 description 1
- 238000012360 testing method Methods 0.000 description 1
- 239000003053 toxin Substances 0.000 description 1
- 231100000765 toxin Toxicity 0.000 description 1
- 238000009966 trimming Methods 0.000 description 1
- 230000002792 vascular Effects 0.000 description 1
- 238000009423 ventilation Methods 0.000 description 1
- 238000005303 weighing 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
Landscapes
- Life Sciences & Earth Sciences (AREA)
- Developmental Biology & Embryology (AREA)
- Engineering & Computer Science (AREA)
- Biotechnology (AREA)
- Cell Biology (AREA)
- Botany (AREA)
- Environmental Sciences (AREA)
- Cultivation Of Plants (AREA)
- Breeding Of Plants And Reproduction By Means Of Culturing (AREA)
Abstract
The invention relates to the technical field of tissue culture and rapid propagation, in particular to a tissue culture seedling method for stem segments of momordica grosvenori. The method comprises the following steps: step 1, cutting young branches of momordica grosvenori into small segments with axillary buds, and sterilizing to obtain sterilized stem segments with axillary buds; step 2, inoculating the stem segment with the axillary bud into a bud induction culture medium, culturing, and inducing adventitious buds; step 3, cutting the induced adventitious buds, inoculating the adventitious buds into a bud multiplication culture medium, and culturing to obtain buds; step 4, transferring the buds to a rooting culture medium, and culturing to obtain tissue culture seedlings; and 5, hardening seedlings of the tissue culture seedlings in a hardening matrix, and then transplanting to obtain the momordica grosvenori seedlings. The invention takes the stem segments of the momordica grosvenori as explants, improves proliferation efficiency by optimizing a tissue culture rapid propagation method, establishes an in-vitro regeneration cultivation system of the high-efficiency mature momordica grosvenori, and provides theoretical basis for industrial and standardized production of high-quality momordica grosvenori seedlings.
Description
Technical Field
The invention relates to the technical field of tissue culture and rapid propagation, in particular to a tissue culture seedling method for stem segments of momordica grosvenori.
Background
Siraitia grosvenorii (Siraitia grosvenorii) belongs to the family Cucurbitaceae, perennial vine. The fructus momordicae flowers and fruits in summer and autumn, and the fruits are rich in nutrition, so that the fructus momordicae is not only a novel functional sweetener, but also a medical and edible Chinese medicinal material with multiple functions. At present, the demand of the medicine industry for momordica grosvenori is gradually increased, and food, functional food and special medical food production enterprises which take the momordica grosvenori extract as raw materials also rapidly rise, so that the development of the momordica grosvenori planting industry is driven.
The propagation mode of the momordica grosvenori seedlings mainly adopts cuttage or vine pressing, but the propagation coefficient of asexual propagation is low, and the propagation coefficient is increased along with the increase of propagation algebra, so that the variety is degenerated, toxin is accumulated, virus diseases are increased, and the yield is greatly reduced.
Disclosure of Invention
The invention aims to provide a tissue culture seedling method for momordica grosvenori stem segments, which can be used for rapidly obtaining high-quality seedlings of momordica grosvenori, solving the problem that the traditional seedling is limited by qualified propagation materials, and simultaneously providing technical support for sustainable development of the momordica grosvenori industry.
In order to achieve the above object, the present invention provides the following solutions:
according to one of the technical schemes, the tissue culture seedling method of the momordica grosvenori stem segment comprises the following steps:
step 1, cutting young branches of momordica grosvenori into small segments with axillary buds, and sterilizing to obtain sterilized stem segments with axillary buds;
step 2, inoculating the disinfected stem section with the axillary buds into a bud induction culture medium, culturing, and inducing adventitious buds;
step 3, cutting the induced adventitious buds, inoculating the adventitious buds into a bud multiplication culture medium, and culturing to obtain buds;
step 4, transferring the buds to a rooting culture medium, and culturing to obtain tissue culture seedlings;
and 5, hardening seedlings of the tissue culture seedlings in a hardening matrix, and then transplanting to obtain the momordica grosvenori seedlings.
Further, in the step 1, before the young branches of the momordica grosvenori are cut into small segments with axillary buds, the young branches of the momordica grosvenori are soaked in 0.05% potassium permanganate solution for 25min and then washed by tap water; the disinfection is specifically as follows: washing with sterile water, sterilizing with 0.1% mercuric chloride for 8min, washing with sterile water, sterilizing with 75% ethanol for 30s, and washing with sterile water.
Further, the composition of the basal medium of the bud induction medium in the step 2 and the basal medium of the bud proliferation medium in the step 3 are MS+25g/L sucrose+7 g/L agar+0.5 g/L Huabao No. 1.
Further, in the step 2, the bud induction medium contains 1.8-2.2mg/L of luffa extract or 6-benzylaminopurine; the loofah sponge extract or 6-benzylaminopurine content higher or lower than the above range is not favorable for bud induction culture.
The bud multiplication medium in step 3 contains 0.8-1.2mg/L of luffa extract and 0.1mg/L of alpha-naphthylacetic acid, or 0.8-1.2mg/L of 6-benzylaminopurine and 0.1mg/L of alpha-naphthylacetic acid. The loofah extract or 6-benzylaminopurine, alpha-naphthylacetic acid content above or below the above range is not favorable for bud proliferation.
Further preferably, in step 2, the sprout induction medium contains 1.8-2.2mg/L of retinervus Luffae fructus extract; the bud proliferation medium in the step 3 contains 0.8-1.2mg/L of retinervus Luffae fructus extract and 0.1mg/L of alpha-naphthylacetic acid.
Further, in step 2, the composition of the bud induction medium is: MS+25g/L sucrose+7 g/L agar+0.5 g/L Huabao No. 1+2.0 mg/L retinervus Luffae fructus extract.
Further, in step 2, the culturing is specifically: the temperature is 26+/-2 ℃, the light is irradiated for 10-12 hours every day, the light intensity is 1000lx, and the culture period is 21d.
Further, in step 3, the composition of the bud proliferation medium is: MS+25g/L sucrose+7 g/L agar+0.5 g/L Huabao No. 1+1.0 mg/L retinervus Luffae fructus extract+0.1 mg/L alpha-naphthylacetic acid.
Further, in step 3, the culturing is specifically: the temperature is 26+/-2 ℃, the light is irradiated for 10-12 hours every day, the light intensity is 1000lx, and the culture period is 21d.
Further, the preparation method of the loofah sponge extract comprises the following steps:
mixing retinervus Luffae fructus and water at volume ratio of 1:1, boiling with strong fire, decocting with slow fire for 15-20min, filtering to obtain first filtrate and residue, mixing residue with water at volume ratio of 1:1, boiling with strong fire, decocting with slow fire for 15-20min, and filtering to obtain second filtrate; combining the first filtrate and the second filtrate, and concentrating under reduced pressure to 1/10-1/8 of the original volume to obtain retinervus Luffae fructus extract.
The retinervus Luffae fructus is the vascular bundle of dried mature fruit of Luffa cylindrica belonging to Cucurbitaceae, and has porous structure. In the preparation of the retinervus Luffae fructus extract, water is preferably used in an amount to completely submerge retinervus Luffae fructus or residue, and therefore the amount of water is limited to the above-described amount.
Further, in the step 4, the rooting medium comprises MS+25g/L sucrose+7 g/L agar+0.5 g/L Huabao No. 1+0.2 g/L carbon powder+0.1 mg/L indolebutyric acid+0.1 mg/L alpha-naphthylacetic acid; the culture is specifically as follows: the temperature is 26+/-2 ℃, the illumination is carried out for 16 hours every day, the illumination intensity is 1000lx, and the culture period is 3-4 weeks.
Further, in step 5, the seedling hardening matrix comprises the following components: vermiculite to perlite to moss volume ratio = 2:1:1.
The invention discloses the following technical effects:
the invention utilizes the tissue culture rapid propagation technology to induce the stem bud multiplication of the momordica grosvenori, and can cultivate high-quality momordica grosvenori seedlings after strengthening seedlings and rooting, so that a large number of high-quality seedlings can be cultivated in a short time, the popularization speed is increased, and the method is a fundamental way for solving the problem of momordica grosvenori virus diseases.
The invention takes the stem segments of the momordica grosvenori as explants, improves proliferation efficiency by optimizing a tissue culture rapid propagation method, establishes an in-vitro regeneration cultivation system of the high-efficiency mature momordica grosvenori, and provides theoretical basis for industrial and standardized production of high-quality momordica grosvenori seedlings.
The loofah sponge is generally used as a dish washing or traditional Chinese medicine, and the loofah sponge extract is creatively used for tissue culture seedlings of the momordica grosvenori, so that the loofah sponge has a good promoting effect on the induction of the buds of the momordica grosvenori. The loofah sponge extract-added bud induction culture medium and bud multiplication culture medium are used in the bud induction and bud multiplication stage of the momordica grosvenori tissue culture seedling, so that more high-quality seedlings can be cultivated in a short time, and the method has important significance for an in-vitro regeneration cultivation system of the momordica grosvenori.
Drawings
In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings that are needed in the embodiments will be briefly described below, and it is obvious that the drawings in the following description are only some embodiments of the present invention, and other drawings may be obtained according to these drawings without inventive effort for a person skilled in the art.
FIG. 1 shows the bud induction of examples 1-3 of the present invention in different bud induction media; wherein A is example 2, B is example 1, and C is example 3.
FIG. 2 shows the bud proliferation of examples 1, 4-5 of the present invention in different bud proliferation media; wherein A is example 4, B is example 1, and C is example 5.
FIG. 3 shows rooting conditions of examples 1, 6-7 according to the invention in different rooting media; wherein A is example 6, B is example 1, and C is example 7.
FIG. 4 is a schematic view of seedling hardening and transplanting in example 1 of the present invention.
Detailed Description
Various exemplary embodiments of the invention will now be described in detail, which should not be considered as limiting the invention, but rather as more detailed descriptions of certain aspects, features and embodiments of the invention.
It is to be understood that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting of the invention. In addition, for numerical ranges in this disclosure, it is understood that each intermediate value between the upper and lower limits of the ranges is also specifically disclosed. Every smaller range between any stated value or stated range, and any other stated value or intermediate value within the stated range, is also encompassed within the invention. The upper and lower limits of these smaller ranges may independently be included or excluded in the range.
Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Although only preferred methods and materials are described herein, any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present invention. All documents mentioned in this specification are incorporated by reference for the purpose of disclosing and describing the methods and/or materials associated with the documents. In case of conflict with any incorporated document, the present specification will control.
It will be apparent to those skilled in the art that various modifications and variations can be made in the specific embodiments of the invention described herein without departing from the scope or spirit of the invention. Other embodiments will be apparent to those skilled in the art from consideration of the specification of the present invention. The specification and examples of the present invention are exemplary only.
As used herein, the terms "comprising," "including," "having," "containing," and the like are intended to be inclusive and mean an inclusion, but not limited to.
The raw materials used in the examples of the present invention, unless otherwise specified, were all available commercially.
"Huabao No. 1" used in the examples of the present invention is purchased from Laibaote organism, and the ratio of the components is: and N is P, K=7:6:19, and the water-soluble instant fertilizer.
The MS medium used in the examples of the present invention was purchased from Soy palettes.
The specific preparation method of the basic culture medium MS+25g/L sucrose+7g/L agar+0.5g/L Huabao No. 1 used in the embodiment of the invention comprises the following steps: weighing 2.3g of MS culture medium, 25g of sucrose, 7g of agar powder and 0.5g of Huabao No. 1, adding 1000mL of distilled water, boiling for dissolution, adjusting the pH value to 5.7-5.9, and sterilizing under high pressure for standby.
The loofah sponge extract used in the embodiment of the invention is prepared by the following steps:
adding retinervus Luffae fructus into water (the volume ratio of retinervus Luffae fructus to water is 1:1), boiling with strong fire, decocting with slow fire for 20min, filtering to obtain first filtrate and residue, adding the residue into water (the volume ratio of residue to water is 1:1), boiling with strong fire, decocting with slow fire for 20min, and filtering to obtain second filtrate; combining the first filtrate and the second filtrate, and concentrating under reduced pressure to 1/10 of the original volume to obtain retinervus Luffae fructus extract.
Example 1
Step 1, explant disinfection treatment: washing young branches of momordica grosvenori with clear water, soaking the young branches in 0.05% potassium permanganate solution for 25min, washing the young branches with tap water, placing the washed young branches into an ultra-clean workbench, shearing the young branches of momordica grosvenori into small sections with axillary buds of 0.5cm, washing the small sections with sterile water for 3 times, disinfecting the small sections with the axillary buds with 0.1% mercuric chloride for 8min, washing the small sections with sterile water for 3-5 times, disinfecting the small sections with alcohol with the volume concentration of 75% for 30s, washing the small sections with sterile water for 3 times, placing the small sections on sterile filter paper, and absorbing surface moisture to obtain disinfected stem sections with the axillary buds.
Step 2, induction culture of buds: inoculating the sterilized stem with axillary bud into a bud induction culture medium, wherein the composition of the bud induction culture medium is MS+25g/L sucrose+7 g/L agar+0.5 g/L Huabao No. 1+2.0 mg/L luffa extract; the culture conditions are that the culture is carried out in culture room bottles at 26+/-2 ℃, the number of inoculated tender stems is 3 per bottle, the illumination is 12 hours per day, the illumination intensity is 1000lx, and the culture period is 21d. The survival rate of the axillary buds at this step was 81%, and the bud induction was shown in FIG. 1B.
Step 3, subculture proliferation: the adventitious buds induced by cutting are inoculated into a bud multiplication culture medium, wherein the composition of the bud multiplication culture medium is MS+25g/L sucrose+7 g/L agar+0.5 g/L flower bud No. 1+1.0 mg/L luffa extract+0.1 mg/LNAA (alpha-naphthylacetic acid), and the culture conditions are as follows: culturing in 26+ -2deg.C culture room bottle, inoculating 2 tender stems per bottle, and illuminating for 12 hr each day with illumination intensity 1000lx for 21d culture period. The statistical bud multiplication factor was 2.77, and the bud multiplication situation is shown in FIG. 2B.
Step 4, rooting culture: transferring the strong bud seedlings obtained in the step 3 into a rooting culture medium, wherein the rooting culture medium comprises the following components: MS+25g/L sucrose+7 g/L agar+0.5 g/L Huabao No. 1+0.2 g/L carbon powder+0.1 mg/LIBA (indolebutyric acid) +0.1mg/LNAA (. Alpha. -naphthylacetic acid), the culture conditions being: culturing in 26+ -2deg.C culture room bottles, inoculating 1 tender stem per bottle, and illuminating for 16 hr per day with illumination intensity of 1000lx. The culture period is 3 weeks (3-4 weeks are all available). Rooting is shown in fig. 3B.
Step 5, hardening seedlings and transplanting: and (3) opening a bottle cap when 4-5 main roots and 3-5 new leaves grow out of the tissue culture seedling in the step (4) and the height is about 10cm, taking out the rooted sterile tissue culture seedling after hardening the seedling in a greenhouse for 2-3 days, cleaning a root culture medium with sterile water, properly trimming root length and leaves, inoculating the tissue culture seedling in a seedling hardening matrix, and hardening the seedling in a greenhouse under natural illumination and natural temperature. The matrix combination is as follows: vermiculite to perlite to moss volume ratio = 2:1:1. After transplanting, the ventilation and moisture retention are enhanced. And after 4 weeks, observing and counting the growth condition of the transplanted seedlings, and obtaining the survival rate of 100%. A schematic diagram of seedling hardening and transplanting in this example is shown in FIG. 4.
Example 2
The only difference from example 1 is that in step 2, the composition of the shoot induction medium was MS+25g/L sucrose+7 g/L agar+0.5 g/L Huabao No. 1+2.0 mg/L6-BA (6-benzylaminopurine). Results: bud induction is shown in FIG. 1A.
Example 3
The only difference from example 1 is that in step 2, the composition of the shoot induction medium was MS+25g/L sucrose+7 g/L agar+0.5 g/L Huabao No. 1+3.0 mg/L6-BA (6-benzylaminopurine). Results: bud induction is shown in figure 1, C.
Example 4
The only difference from example 1 is that in step 3, the composition of the shoot proliferation medium was MS+25g/L sucrose+7 g/L agar+0.5 g/L Huabao No. 1+1.0 mg/L6-BA (6-benzylaminopurine) +0.1mg/LNAA (. Alpha. -naphthylacetic acid). Results: bud proliferation is shown in FIG. 2A.
Example 5
The only difference from example 1 is that in step 3, the composition of the shoot proliferation medium was MS+25g/L sucrose+7 g/L agar+0.5 g/L Huabao No. 1+1.0 mg/L retinervus luffae fructus extract+0.12 mg/LNAA (. Alpha. -naphthylacetic acid). Results: bud proliferation is shown in FIG. 2C.
Example 6
The only difference from example 1 is that in step 4, the composition of the rooting medium is: MS+25g/L sucrose+7 g/L agar+0.5 g/L Huabao No. 1+0.2 g/L carbon powder+0.1 mg/LIBA (indolebutyric acid) +0.05mg/LNAA (. Alpha. -naphthylacetic acid). Results: rooting is shown in figure 3 a.
Example 7
The only difference from example 1 is that in step 4, the composition of the rooting medium is: MS+25g/L sucrose+7 g/L agar+0.5 g/L Huabao No. 1+0.2 g/L carbon powder+0.1 mg/LIBA (indolebutyric acid) +0.15mg/LNAA (. Alpha. -naphthylacetic acid). Results: rooting is shown in figure 3C.
Example 8
The only difference from example 1 is that the matrix combination in step 5 is: vermiculite to perlite volume ratio = 2:1. Results: the survival rate of the transplanted seedlings is 62 percent
Example 9
The only difference from example 1 is that the matrix combination in step 5 is: vermiculite: perlite: moss: bark = 1:1:1:1. Results: the survival rate of the transplanted seedlings is 73 percent
The invention also tries to apply the luffa extract to the rooting culture stage of the tissue culture seedling of the fructus momordicae, and the rooting promoting effect is not as good as that of the combination of 0.1mg/LIBA (indolebutyric acid) +0.1mg/LNAA (alpha-naphthylacetic acid).
The combination of the vermiculite, the perlite and the moss (the surface covered by the moss is used for moisturizing) matrix can achieve better hydrophobic and water-retaining capacities, and is more beneficial to the growth of the momordica grosvenori compared with the combination of the vermiculite, the perlite and the moss.
The invention establishes a simple, safe and efficient momordica grosvenori tissue culture seedling technical system through a tissue culture means, can provide theoretical basis for industrial production of high-quality momordica grosvenori seedlings, and also establishes a new way for mass production of momordica grosvenori detoxified seedlings through a tissue culture technology.
The above embodiments are only illustrative of the preferred embodiments of the present invention and are not intended to limit the scope of the present invention, and various modifications and improvements made by those skilled in the art to the technical solutions of the present invention should fall within the protection scope defined by the claims of the present invention without departing from the design spirit of the present invention.
Claims (6)
1. The tissue culture seedling method of the momordica grosvenori stem segment is characterized by comprising the following steps of:
step 1, cutting young branches of momordica grosvenori into small segments with axillary buds, and sterilizing to obtain sterilized stem segments with axillary buds;
step 2, inoculating the disinfected stem section with the axillary buds into a bud induction culture medium, culturing, and inducing adventitious buds;
step 3, cutting the induced adventitious buds, inoculating the adventitious buds into a bud multiplication culture medium, and culturing to obtain buds;
step 4, transferring the buds to a rooting culture medium, and culturing to obtain tissue culture seedlings;
step 5, hardening seedlings of the tissue culture seedlings in a hardening matrix, and then transplanting to obtain momordica grosvenori seedlings;
the composition of the basal medium of the bud induction medium in the step 2 and the bud proliferation medium in the step 3 is MS+25g/L sucrose+7 g/L agar+0.5 g/L Huabao No. 1;
in the step 2, the bud induction culture medium contains 1.8-2.2mg/L of luffa extract; the bud proliferation culture medium in the step 3 contains 0.8-1.2mg/L of luffa extract and 0.1mg/L of alpha-naphthylacetic acid;
the preparation method of the loofah sponge extract comprises the following steps:
mixing retinervus Luffae fructus and water at volume ratio of 1:1, boiling with strong fire, decocting with slow fire for 15-20min, filtering to obtain first filtrate and residue, mixing residue with water at volume ratio of 1:1, boiling with strong fire, decocting with slow fire for 15-20min, and filtering to obtain second filtrate; combining the first filtrate and the second filtrate, and concentrating under reduced pressure to 1/10-1/8 of the original volume to obtain retinervus Luffae fructus extract;
in the step 4, the rooting culture medium comprises MS+25g/L sucrose+7 g/L agar+0.5 g/L Huabao No. 1+0.2 g/L carbon powder+0.1 mg/L indolebutyric acid+0.1 mg/L alpha-naphthylacetic acid; the culture is specifically as follows: the temperature is 26+/-2 ℃, the illumination is carried out for 16 hours every day, the illumination intensity is 1000lx, and the culture period is 3-4 weeks.
2. The method for tissue culture of stem segments of momordica grosvenori according to claim 1, wherein in the step 2, the composition of the bud induction medium is: MS+25g/L sucrose+7 g/L agar+0.5 g/L Huabao No. 1+2.0 mg/L retinervus Luffae fructus extract.
3. The method for tissue culture seedling of momordica grosvenori stem segments according to claim 1, wherein in the step 2, the culturing is specifically: the temperature is 26+/-2 ℃, the light is irradiated for 10-12 hours every day, the light intensity is 1000lx, and the culture period is 21d.
4. The method for tissue culture seedling of stem segments of momordica grosvenori according to claim 1, wherein in the step 3, the composition of the bud proliferation medium is as follows: MS+25g/L sucrose+7 g/L agar+0.5 g/L Huabao No. 1+1.0 mg/L retinervus Luffae fructus extract+0.1. 0.1mg/L alpha-naphthylacetic acid.
5. The method for tissue culture seedling of momordica grosvenori stem segments according to claim 1, wherein in the step 3, the culturing is specifically: the temperature is 26+/-2 ℃, the illumination is 10-12h per day, the illumination intensity is 1000lx, and the culture period is 21d.
6. The method for tissue culture seedling of momordica grosvenori stem segments according to claim 1, wherein in the step 5, the seedling hardening matrix comprises the following components: vermiculite to perlite to moss volume ratio = 2:1:1.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202310426135.3A CN116458428B (en) | 2023-04-20 | 2023-04-20 | Tissue culture seedling method for momordica grosvenori stem segments |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202310426135.3A CN116458428B (en) | 2023-04-20 | 2023-04-20 | Tissue culture seedling method for momordica grosvenori stem segments |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN116458428A CN116458428A (en) | 2023-07-21 |
| CN116458428B true CN116458428B (en) | 2023-12-22 |
Family
ID=87180280
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202310426135.3A Active CN116458428B (en) | 2023-04-20 | 2023-04-20 | Tissue culture seedling method for momordica grosvenori stem segments |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN116458428B (en) |
Families Citing this family (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN118216323A (en) * | 2024-03-21 | 2024-06-21 | 湖南农夫哥农业开发有限公司 | Root-plug seedling method for lonicera macranthoides |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2021170618A1 (en) * | 2020-02-25 | 2021-09-02 | Clariant International Ltd | Compositions comprising luffa cylindrica root extract |
| CN113661923A (en) * | 2021-06-25 | 2021-11-19 | 南宁市农业科学研究所 | Rapid propagation method of siraitia grosvenorii seedlings |
Family Cites Families (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| DE102018100485A1 (en) * | 2018-01-10 | 2019-07-11 | RoBoTec PTC GmbH | Nutrient medium for the automated production of plants |
-
2023
- 2023-04-20 CN CN202310426135.3A patent/CN116458428B/en active Active
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2021170618A1 (en) * | 2020-02-25 | 2021-09-02 | Clariant International Ltd | Compositions comprising luffa cylindrica root extract |
| CN113661923A (en) * | 2021-06-25 | 2021-11-19 | 南宁市农业科学研究所 | Rapid propagation method of siraitia grosvenorii seedlings |
Non-Patent Citations (3)
| Title |
|---|
| 田间"碧秀"苦瓜茎段离体启动培养的研究;刘玉晗;王永清;秦红梅;;北方园艺(第11期);148-150 * |
| 罗汉果高效组培快繁技术的探究及优化;谭秀梅;王晓英;田启建;夏勉;刘世彪;;药物生物技术(第06期);518-521 * |
| 谭秀梅 ; 王晓英 ; 田启建 ; 夏勉 ; 刘世彪 ; .罗汉果高效组培快繁技术的探究及优化.药物生物技术.2013,(第06期),518-521. * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN116458428A (en) | 2023-07-21 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN108293878B (en) | Tissue culture seedling raising method for trichosanthes kirilowii Maxim tender leaves | |
| CN101507415B (en) | In-vitro culture method of antlerpilose grass | |
| CN116458428B (en) | Tissue culture seedling method for momordica grosvenori stem segments | |
| CN110810250B (en) | Method for rapid propagation in tissue culture of date palm | |
| CN108077071B (en) | Culture medium for culturing vitex agnus-castus tissue and rapid propagation method | |
| CN111557138A (en) | Treatment method for improving tissue culture germination of myrtle seeds and reducing pollution rate | |
| CN113875585A (en) | Method for in-vitro rapid propagation and seedling raising of roxburgh rose | |
| CN112655553A (en) | Rapid sterile short-shoot propagation method for Orthosiphon aristatus | |
| CN108651275A (en) | A kind of method of quickly breeding bletilla striata seedling under natural light | |
| CN115843688A (en) | Tissue culture seedling raising method by utilizing stem tip grafting of gardenia | |
| CN110612905B (en) | Tissue culture rapid propagation method of dracocephalum plants and application thereof | |
| CN115152629A (en) | Raspberry tissue culture method | |
| CN109258463B (en) | Vegetative propagation method of paphiopedilum armeniacum | |
| CN109997692B (en) | Cyclocarya paliurus callus induction and subculture multiplication culture medium and culture method thereof | |
| CN112806262A (en) | Tissue culture efficient propagation method for betula halophila | |
| CN115885847B (en) | Culture medium for tissue culture and rapid propagation of sightseeing wood, application of culture medium and method for tissue culture and rapid propagation of sightseeing wood | |
| CN115997684B (en) | Culture medium and culture method for tissue culture and seedling raising of cercis chinensis | |
| CN112753579B (en) | In-vitro culture and plant regeneration method for leaves of Chinese medicinal herb chlorophytum comosum | |
| CN109169281A (en) | A kind of rose preserving seed method based on tissue cultures | |
| CN116369203B (en) | Lycoris plant floret regeneration medium and floret regeneration method | |
| CN108739381A (en) | A kind of method for tissue culture of Garbo fruit | |
| CN114208680B (en) | Tissue culture and rapid propagation method for sedum formosanum | |
| CN114190275B (en) | Tissue culture rapid propagation method for one-step seedling formation of rubia yunnanensis | |
| CN110521595B (en) | Method for promoting stem elongation in vitro culture of oriental cherry | |
| CN1465229A (en) | Method for cultivating Aralia elata seen tissue |
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 | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |