CN117859652A - Fructus phyllanthi tissue culture rapid propagation seedling method and application - Google Patents
Fructus phyllanthi tissue culture rapid propagation seedling method and application Download PDFInfo
- Publication number
- CN117859652A CN117859652A CN202410278531.0A CN202410278531A CN117859652A CN 117859652 A CN117859652 A CN 117859652A CN 202410278531 A CN202410278531 A CN 202410278531A CN 117859652 A CN117859652 A CN 117859652A
- Authority
- CN
- China
- Prior art keywords
- culture
- rooting
- culture medium
- days
- fructus phyllanthi
- 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
- 239000009609 fructus phyllanthi Substances 0.000 title claims abstract description 28
- 238000000034 method Methods 0.000 title claims abstract description 20
- 230000035755 proliferation Effects 0.000 claims abstract description 22
- 230000001954 sterilising effect Effects 0.000 claims abstract description 22
- 230000006698 induction Effects 0.000 claims abstract description 21
- 239000001963 growth medium Substances 0.000 claims abstract description 19
- 238000004659 sterilization and disinfection Methods 0.000 claims abstract description 13
- 238000012258 culturing Methods 0.000 claims abstract description 12
- 229960002523 mercuric chloride Drugs 0.000 claims abstract description 10
- LWJROJCJINYWOX-UHFFFAOYSA-L mercury dichloride Chemical compound Cl[Hg]Cl LWJROJCJINYWOX-UHFFFAOYSA-L 0.000 claims abstract description 10
- 239000012883 rooting culture medium Substances 0.000 claims abstract description 8
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims abstract description 7
- ZKQDCIXGCQPQNV-UHFFFAOYSA-N Calcium hypochlorite Chemical class [Ca+2].Cl[O-].Cl[O-] ZKQDCIXGCQPQNV-UHFFFAOYSA-N 0.000 claims abstract description 6
- 238000004140 cleaning Methods 0.000 claims abstract description 6
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 claims description 21
- YGSDEFSMJLZEOE-UHFFFAOYSA-N salicylic acid Chemical compound OC(=O)C1=CC=CC=C1O YGSDEFSMJLZEOE-UHFFFAOYSA-N 0.000 claims description 20
- 238000005286 illumination Methods 0.000 claims description 15
- 239000008223 sterile water Substances 0.000 claims description 15
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 15
- FJKROLUGYXJWQN-UHFFFAOYSA-N papa-hydroxy-benzoic acid Natural products OC(=O)C1=CC=C(O)C=C1 FJKROLUGYXJWQN-UHFFFAOYSA-N 0.000 claims description 10
- 229960004889 salicylic acid Drugs 0.000 claims description 10
- 238000005507 spraying Methods 0.000 claims description 10
- XSQUKJJJFZCRTK-UHFFFAOYSA-N Urea Chemical compound NC(N)=O XSQUKJJJFZCRTK-UHFFFAOYSA-N 0.000 claims description 9
- 239000004202 carbamide Substances 0.000 claims description 9
- 239000011159 matrix material Substances 0.000 claims description 9
- 235000015489 Emblica officinalis Nutrition 0.000 claims description 8
- 230000000249 desinfective effect Effects 0.000 claims description 8
- 239000001888 Peptone Substances 0.000 claims description 7
- 108010080698 Peptones Proteins 0.000 claims description 7
- 235000019319 peptone Nutrition 0.000 claims description 7
- 229910052799 carbon Inorganic materials 0.000 claims description 5
- 238000009472 formulation Methods 0.000 claims description 4
- 239000000203 mixture Substances 0.000 claims description 4
- 239000012882 rooting medium Substances 0.000 claims description 2
- 239000002609 medium Substances 0.000 claims 2
- 240000009120 Phyllanthus emblica Species 0.000 claims 1
- 239000003610 charcoal Substances 0.000 claims 1
- 238000004161 plant tissue culture Methods 0.000 abstract description 2
- 244000119298 Emblica officinalis Species 0.000 description 7
- 230000000052 comparative effect Effects 0.000 description 6
- 235000013162 Cocos nucifera Nutrition 0.000 description 4
- 244000060011 Cocos nucifera Species 0.000 description 4
- 241000196324 Embryophyta Species 0.000 description 4
- 235000013399 edible fruits Nutrition 0.000 description 4
- 230000000694 effects Effects 0.000 description 4
- 239000010903 husk Substances 0.000 description 4
- 239000003415 peat Substances 0.000 description 4
- 235000019362 perlite Nutrition 0.000 description 4
- 239000010451 perlite Substances 0.000 description 4
- 239000002689 soil Substances 0.000 description 4
- 230000004083 survival effect Effects 0.000 description 4
- 238000004017 vitrification Methods 0.000 description 4
- 238000009395 breeding Methods 0.000 description 3
- 230000001488 breeding effect Effects 0.000 description 3
- 238000011109 contamination Methods 0.000 description 2
- 238000005520 cutting process Methods 0.000 description 2
- 238000011161 development Methods 0.000 description 2
- 230000018109 developmental process Effects 0.000 description 2
- 239000003814 drug Substances 0.000 description 2
- 235000012055 fruits and vegetables Nutrition 0.000 description 2
- 238000012545 processing Methods 0.000 description 2
- 241000221017 Euphorbiaceae Species 0.000 description 1
- 206010062717 Increased upper airway secretion Diseases 0.000 description 1
- 206010028980 Neoplasm Diseases 0.000 description 1
- 206010068319 Oropharyngeal pain Diseases 0.000 description 1
- 201000007100 Pharyngitis Diseases 0.000 description 1
- 206010039424 Salivary hypersecretion Diseases 0.000 description 1
- GAMYVSCDDLXAQW-AOIWZFSPSA-N Thermopsosid Natural products O(C)c1c(O)ccc(C=2Oc3c(c(O)cc(O[C@H]4[C@H](O)[C@@H](O)[C@H](O)[C@H](CO)O4)c3)C(=O)C=2)c1 GAMYVSCDDLXAQW-AOIWZFSPSA-N 0.000 description 1
- 230000032683 aging Effects 0.000 description 1
- 150000001413 amino acids Chemical class 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 235000013361 beverage Nutrition 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 229930003944 flavone Natural products 0.000 description 1
- 150000002212 flavone derivatives Chemical class 0.000 description 1
- 235000011949 flavones Nutrition 0.000 description 1
- 239000000796 flavoring agent Substances 0.000 description 1
- 235000019634 flavors Nutrition 0.000 description 1
- 235000013305 food Nutrition 0.000 description 1
- 235000011389 fruit/vegetable juice Nutrition 0.000 description 1
- 230000010196 hermaphroditism Effects 0.000 description 1
- 238000011081 inoculation Methods 0.000 description 1
- 239000007937 lozenge Substances 0.000 description 1
- 210000004072 lung Anatomy 0.000 description 1
- 230000014759 maintenance of location Effects 0.000 description 1
- 235000013372 meat Nutrition 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 235000016709 nutrition Nutrition 0.000 description 1
- 230000035764 nutrition Effects 0.000 description 1
- 230000003647 oxidation Effects 0.000 description 1
- 238000007254 oxidation reaction Methods 0.000 description 1
- 208000026435 phlegm Diseases 0.000 description 1
- 230000002035 prolonged effect Effects 0.000 description 1
- 230000001737 promoting effect Effects 0.000 description 1
- 238000010791 quenching Methods 0.000 description 1
- 230000000171 quenching effect Effects 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 208000026451 salivation Diseases 0.000 description 1
- 230000002000 scavenging effect Effects 0.000 description 1
- 238000002791 soaking Methods 0.000 description 1
- 235000018553 tannin Nutrition 0.000 description 1
- 229920001864 tannin Polymers 0.000 description 1
- 239000001648 tannin Substances 0.000 description 1
- 230000035922 thirst Effects 0.000 description 1
- VHBFFQKBGNRLFZ-UHFFFAOYSA-N vitamin p Natural products O1C2=CC=CC=C2C(=O)C=C1C1=CC=CC=C1 VHBFFQKBGNRLFZ-UHFFFAOYSA-N 0.000 description 1
Landscapes
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
The invention relates to the technical field of plant tissue culture, in particular to a fructus phyllanthi tissue culture rapid propagation seedling method and application, and the method comprises the following steps: cleaning immature stem segments at the top of the non-lignified fructus phyllanthi, and sequentially sterilizing with 75% alcohol, saturated bleaching powder solution and 0.15% mercuric chloride solution; inoculating the sterilized explant into the primary induction culture medium for culturing for 25-35 days; inoculating the primary induced cluster buds into a secondary proliferation culture medium for culturing for 30-40 days; and inoculating the cluster buds subjected to the secondary multiplication culture into a rooting culture medium for culturing for 30-40 days. The invention changes the concentration of the conventional mercuric chloride by 0.1% into 0.15%, can control the disinfection pollution rate of the explant within 15% and the death rate within 20%, and can minimize the occurrence or influence of browning by assisting with the primary induction culture medium, the primary induction rate can be up to more than 62%, the proliferation coefficient can be up to about 5.5, and the rooting rate can be up to more than 95%.
Description
Technical Field
The invention relates to the technical field of plant tissue culture, in particular to a method for quickly breeding seedlings by utilizing phyllanthus emblica tissue culture and application thereof.
Background
Fructus phyllanthiPhyllanthus emblica Linn .) The Yunnan phyllanthus emblica is the fruit of phyllanthus emblica which is a falling leaf shrub or arbor plant of Euphorbiaceae. PlantingFlowering in spring, small flowers, single yellow, hermaphrodite strains. The meat quality of the fruits, the oblate spheroids, and the fruits are red when cooked. Is produced in the tropical region of Asia, and is common in the south of China. Is an important plant resource for both medicine and food. The fructus phyllanthi juice has unique flavor and rich nutrition, contains amino acids and microelements, and more importantly, contains V c High content and V of fructus Phyllanthi after processing due to protection of flavone and tannin c The retention rate of the fruit and vegetable is still up to 72 percent and is far higher than that of other fruits and vegetables. The fruit is used as medicine, and has effects of clearing heat, relieving sore throat, moistening lung, resolving phlegm, promoting salivation, and quenching thirst. Modern researches have led to the effects of resisting oxidation, scavenging free radicals, resisting tumor and resisting aging.
With the development of modern processing technology, products (beverages, wine soaking, lozenge and the like) related to fructus phyllanthi are greatly developed and achieve positive market effects, so that the demand of fructus phyllanthi is increased, and artificial planting of fructus phyllanthi has rapidly developed in many regions of Yunnan. The traditional planting mode requires technicians to directly select seeds in the field, and seedlings of fructus phyllanthi are obtained through modes of seed propagation, cutting, grafting and the like after the selected varieties are collected. However, the seedling breeding of fructus phyllanthi by means of seed, cutting, grafting and the like has the problems of long breeding period, high death rate and the like. The publication number is: the Chinese patent of CN107960328A discloses a method for tissue culture of phyllanthus emblica, which realizes the aim of rapid propagation of phyllanthus emblica, but aims at the problems of low propagation coefficient, low rooting rate and the like of phyllanthus emblica varieties in the actual operation process.
Disclosure of Invention
The invention aims at solving the problems pointed out in the background art, and provides a method for tissue culture and rapid propagation of fructus phyllanthi and application thereof, so that the rapid propagation of fructus phyllanthi is realized, and the development requirement of fructus phyllanthi industry is met.
In order to achieve the above purpose, the invention adopts the following technical scheme: a fructus phyllanthi tissue culture rapid propagation seedling method comprises the following steps:
(1) Explant sterilization: taking a young stem section at the top of a non-lignified wild fructus phyllanthi, cleaning, disinfecting with 75% alcohol for 30-40 s and sterile water for 3-4 times, disinfecting with saturated bleaching powder solution for 40-50 min and sterile water for 2-3 times, disinfecting with 0.15% mercuric chloride solution for 2-3 min and disinfecting with sterile water for 7-8 times;
(2) Primary induction culture: inoculating the sterilized explant into the primary induction culture medium for culturing for 25-35 days;
the formula of the primary induction culture medium is as follows: WPM+NAA 0.02-0.05 mg/L+ZT1.0-1.2 mg/L+6-BA 3.0-4.0 mg/L+GA 3 0.1-0.2 mg/L+0.8-1.3 g/L of active carbon;
(3) And (3) subculturing and proliferation: inoculating the primary induced cluster buds into a secondary proliferation culture medium for culturing for 30-40 days;
the formula of the secondary proliferation culture medium is as follows: 0.01 to 0.02mg/L of WPM+NAA+0.8 to 1.0mg/L of TDZ+0.5 to 1.0mg/L of ZT+0.05 to 0.1g/L of urea;
(4) Rooting culture: inoculating cluster buds subjected to secondary multiplication culture into a rooting culture medium for culturing for 30-40 days;
the rooting culture medium comprises the following formula: SH+IBA 0.2-0.4 mg/L+IAA 1.0-1.5 mg/L+PPP 333 0.02-0.05 mg/L + peptone 100-200 mg/L + activated carbon 0.1-0.3 g/L;
(5) Domesticating rooting seedlings: transplanting the tissue culture rooting seedling into rooting matrix (the rooting matrix is peat soil, coconut husk and perlite, the weight ratio of which is 4:6:1), spraying 30-40 mg/L salicylic acid once after transplanting under a greenhouse with a domestication environment of 65-75% shading rate, and spraying 30-40 mg/L salicylic acid once every 5-7 days.
Further, in the steps (2), (3) and (4), the culture conditions are: the temperature is 23+/-2 ℃, the illumination intensity is 2500-3000 Lx, and the illumination is 8-14 hours for light-dark alternate culture.
Further, the formula of the primary induction culture medium is as follows: WPM+NAA 0.04 mg/L+ZT1.1 mg/L+6-BA 3.5mg/L+GA 3 0.15 mg/L+1.0 g/L of activated carbon.
Further, the formula of the secondary proliferation culture medium is as follows: WPM+NAA 0.01mg/L+TDZ 0.9mg/L+ZT 0.8 mg/L+urea 0.08g/L.
Further, the rooting medium comprises the following formula: SH+IBA 0.3mg/L+IAA 1.2mg/L+PPP 333 0.04 mg/L+150 mg/L peptone+0.2 g/L activated carbon.
The beneficial technical effects of the invention are as follows:
1. the method generally adopts 75% alcohol and 0.1% mercuric chloride to carry out different disinfection time combinations when a sterile system is established by tissue culture, but when the explant of fructus phyllanthi is disinfected, the disinfection time of 0.1% mercuric chloride is too short, the pollution rate is extremely high, the sterile system can not be established basically, and when the disinfection time of 0.1% mercuric chloride is prolonged, the disinfection pollution rate is remarkably reduced, but the disinfection death rate is rapidly increased, and even if the explant does not die, cluster buds can not be well induced.
2. In the primary induction culture, NAA, ZT and 6-BA with proper concentration are combined, and GA with certain concentration is added 3 The primary induction rate can reach more than 60 percent, meanwhile, the addition of GA3 can greatly lengthen internode, and the subsequent inoculation operation is convenient.
3. In the invention, NAA, TDZ, ZT with a certain concentration is adopted for matching during the secondary multiplication culture, and urea with a certain concentration is creatively added, so that the multiplication coefficient reaches about 5.5, the color of the multiplied cluster buds is fresh and strong, almost no vitrification phenomenon occurs, and meanwhile, the addition of urea can also inhibit the occurrence of flower buds to a great extent.
4. The invention adopts IBA, IAA and PPP with certain concentration in rooting culture 333 The rooting rate of fructus phyllanthi can reach more than 95%, and meanwhile, the primary and secondary WPM basic culture mediums are changed into SH culture mediums during rooting, and PPP with a certain concentration is added 333 The plant and root system can be made stronger, and the survival rate of the plant and root system is improved when the seedling is further domesticated.
Detailed Description
The technical solutions of the embodiments of the present invention will be clearly and completely described below in conjunction with the embodiments of the present invention, and it is apparent that the described embodiments are only some embodiments of the present invention, not all embodiments. All other embodiments, which can be made by those skilled in the art based on the embodiments of the invention without making any inventive effort, are intended to be within the scope of the invention.
Example 1
A fructus phyllanthi tissue culture rapid propagation seedling method comprises the following steps:
(1) Explant sterilization: taking a non-lignified top tender stem section of a wild fructus phyllanthi in Yunnan, cleaning, sequentially sterilizing with 75% alcohol for 30s and sterile water for 3 times, sterilizing with saturated bleaching powder solution for 40min and sterile water for 3 times, sterilizing with 0.15% mercuric chloride solution for 2min and sterilizing with sterile water for 7 times;
(2) Primary induction culture: the sterilized explants were inoculated in the following formulation: WPM+NAA 0.02 mg/L+ZT1.0 mg/L+6-BA 3.0 mg/L+GA 3 0.1 In a primary induction culture medium of mg/L and 0.8 g/L of activated carbon, the temperature is 23+/-2 ℃, the illumination intensity is 2500Lx, and the light and dark alternate culture is carried out for 10 hours;
(3) And (3) subculturing and proliferation: the primary induced cluster buds are inoculated in the formula: in a secondary proliferation culture medium of WPM+NAA 0.01mg/L+TDZ 0.8mg/L+ZT 0.5 mg/L+urea 0.05g/L, the temperature is 23+/-2 ℃, the illumination intensity is 2500Lx, and the illumination 8 is subjected to light-dark alternate culture for 40 days;
(4) Rooting culture: the cluster buds of the secondary proliferation culture are inoculated in the formula: SH+IBA 0.2mg/L+IAA 1.0mg/L+PPP 333 Culturing in rooting culture medium of 0.02 mg/L+100 mg/L peptone+0.1 g/L active carbon at 23+ -2deg.C under 2500Lx illumination intensity for 8 hr alternately in light and dark for 40 days;
(5) Domestication of root seedlings: transplanting the tissue culture rooting seedling into rooting matrix (the rooting matrix is peat soil, coconut husk and perlite, the weight ratio of which is 4:6:1), spraying 30mg/L salicylic acid once after transplanting under a greenhouse with a domestication environment of 65% shading rate, and spraying 30mg/L salicylic acid once every 5 days.
Example 2
A fructus phyllanthi tissue culture rapid propagation seedling method comprises the following steps:
(1) Explant sterilization: taking a top tender stem section of a non-lignified wild fructus phyllanthi in Yunnan, cleaning, sequentially sterilizing with 75% alcohol for 40s and sterile water for 4 times, sterilizing with saturated bleaching powder solution for 50min and sterile water for 3 times, sterilizing with 0.15% mercuric chloride solution for 3min and sterilizing with sterile water for 8 times;
(2) Primary induction culture: the sterilized explants were inoculated in the following formulation: WPM+NAA 0.05 mg/L+ZT1.2 mg/L+6-BA 4.0mg/L+GA 3 In a primary induction culture medium of 0.2mg/L and 1.3g/L of activated carbon, the temperature is 23+/-2 ℃, the illumination intensity is 3000Lx, and the light and dark alternate culture is carried out for 8 hours;
(3) And (3) subculturing and proliferation: the primary induced cluster buds are inoculated in the formula: in a secondary proliferation culture medium of WPM+NAA 0.02mg/L+TDZ 1.0mg/L+ZT 1.0 mg/L+urea 0.1g/L, the temperature is 23+/-2 ℃, the illumination intensity is 3000Lx, and the light and dark alternate culture is carried out for 8 hours;
(4) Rooting culture: the cluster buds of the secondary proliferation culture are inoculated in the formula: SH+IBA 0.4mg/L+IAA 1.5mg/L+PPP 333 Culturing in rooting culture medium of 0.05 mg/L+200 mg/L peptone+0.3 g/L active carbon at 23+ -2deg.C under 3000Lx illumination intensity for 14 hr alternately under light and dark for 40 days.
(5) Domestication of root seedlings: transplanting the tissue culture rooting seedling into rooting matrix (the rooting matrix is peat soil, coconut husk and perlite, the weight ratio of which is 4:6:1), spraying 40mg/L salicylic acid once after transplanting under a greenhouse with a domestication environment of 75% shading rate, and spraying 40mg/L salicylic acid once every 7 days.
Example 3
A fructus phyllanthi tissue culture rapid propagation seedling method comprises the following steps:
(1) Explant sterilization: taking a top tender stem section of a non-lignified wild fructus phyllanthi in Yunnan, cleaning, sequentially sterilizing with 75% alcohol for 35s and sterile water for 3 times, sterilizing with saturated bleaching powder solution for 45min and sterile water for 3 times, sterilizing with 0.15% mercuric chloride solution for 2.5min and sterilizing with sterile water for 8 times;
(2) First generationInduction culture: the sterilized explants were inoculated in the following formulation: WPM+NAA 0.04 mg/L+ZT1.1 mg/L+6-BA 3.5mg/L+GA 3 In a primary induction culture medium of 0.15mg/L and 1.0g/L of activated carbon, the temperature is 23+/-2 ℃, the illumination intensity is 3000Lx, and the light and dark alternate culture is carried out for 12 hours;
(3) And (3) subculturing and proliferation: the primary induced cluster buds are inoculated in the formula: in a secondary proliferation culture medium of WPM+NAA 0.01mg/L+TDZ 0.9mg/L+ZT 0.8 mg/L+urea 0.08g/L, the temperature is 23+/-2 ℃, the illumination intensity is 2800Lx, and the illumination is 12 hours for light-dark alternate culture for 35 days;
(4) Rooting culture: the cluster buds of the secondary proliferation culture are inoculated in the formula: SH+IBA 0.3mg/L+IAA 1.2mg/L+PPP 333 Culturing in rooting culture medium of 0.04 mg/L+150 mg/L peptone+0.2 g/L active carbon at 23+ -2deg.C under 2800Lx illumination intensity for 12 hr for 35 days.
(5) Domestication of root seedlings: transplanting the tissue culture rooting seedling into rooting matrix (the rooting matrix is peat soil, coconut husk and perlite, the weight ratio of which is 4:6:1), spraying 35mg/L salicylic acid once after transplanting under a greenhouse with a domestication environment of 70% shading rate, and spraying 35mg/L salicylic acid once every 6 days.
Comparative example 1
Taking a top tender stem section of a local wild fructus phyllanthi in Yunnan, which is not lignified, 4cm, according to the Chinese patent invention: example 1 of a method for tissue culture of Emblica officinalis (publication No. CN 107960328A) was conducted.
The results of comparing the sterilizing contamination rate, sterilizing death rate, primary induction rate, proliferation coefficient, flower bud occurrence rate, secondary vitrification rate, rooting rate and survival rate of acclimation for 60 days of the examples 1 to 3 and the comparative example 1 are shown in Table 1 below.
Table 1 comparative results
| Rate of disinfection pollution | Mortality rate of disinfection | First generation induction rate | Proliferation coefficient | Flower bud incidence | Subculture vitrification rate | Rooting rate | Survival rate of acclimatization for 60 days | |
| Example 1 | 12.3% | 17.4% | 71.8% | 5.1 | 0 | 0.47% | 95.7% | 97.7% |
| Example 2 | 13.7% | 15.9% | 65.4% | 5.4 | 0 | 0.36% | 96.3% | 98.2% |
| Example 3 | 11.2% | 10.7% | 79.2% | 6.1 | 0 | 0.43% | 97.4% | 99.3% |
| Comparative example 1 | 40.6% | 38.3% | 60.6% | 4.1 | 3.2% | 6.29% | 83.2% | 87 .6% |
As is clear from Table 1, in examples 1 to 3, the sterilizing contamination rate, sterilizing death rate, proliferation coefficient, flower bud occurrence rate, and subculture vitrification rate were all lower than those of comparative example 1, and the primary induction rate, rooting rate, and survival rate after 60 days of acclimation were higher than those of comparative example 1.
Finally, what should be said is: the above embodiments are only for illustrating the technical aspects of the present invention, and although the present invention has been described in detail with reference to the above embodiments, it should be understood by those skilled in the art that: modifications and equivalents may be made thereto without departing from the spirit and scope of the invention, which is intended to be encompassed by the claims.
Claims (6)
1. The fructus phyllanthi tissue culture rapid propagation seedling method is characterized by comprising the following steps:
(1) Explant sterilization: taking a young stem section at the top of a non-lignified wild fructus phyllanthi, cleaning, disinfecting with 75% alcohol for 30-40 s and sterile water for 3-4 times, disinfecting with saturated bleaching powder solution for 40-50 min and sterile water for 2-3 times, disinfecting with 0.15% mercuric chloride solution for 2-3 min and disinfecting with sterile water for 7-8 times;
(2) Primary induction culture: inoculating the sterilized explant into the primary induction culture medium for culturing for 25-35 days;
the formula of the primary induction culture medium is as follows: WPM+NAA 0.02-0.05 mg/L+ZT1.0-1.2 mg/L+6-BA 3.0-4.0 mg/L+GA 3 0.1-0.2 mg/L+0.8-1.3 g/L of active carbon;
(3) And (3) subculturing and proliferation: inoculating the primary induced cluster buds into a secondary proliferation culture medium for culturing for 30-40 days;
the formula of the secondary proliferation culture medium is as follows: 0.01 to 0.02mg/L of WPM+NAA+0.8 to 1.0mg/L of TDZ+0.5 to 1.0mg/L of ZT+0.05 to 0.1g/L of urea;
(4) Rooting culture: inoculating cluster buds subjected to secondary multiplication culture into a rooting culture medium for culturing for 30-40 days;
the rooting culture medium comprises the following formula: SH+IBA 0.2-0.4 mg/L+IAA 1.0-1.5 mg/L+PPP 333 0.02-0.05 mg/L + peptone 100-200 mg/L + activated carbon 0.1-0.3 g/L;
(5) Domesticating rooting seedlings: transplanting the tissue culture rooting seedling into rooting matrix, spraying 30-40 mg/L salicylic acid once after transplanting under a greenhouse with a domestication environment of 65-75% shading rate, and spraying 30-40 mg/L salicylic acid once every 5-7 days.
2. The method according to claim 1, wherein in the steps (2), (3) and (4), the culture conditions are: the temperature is 23+/-2 ℃, the illumination intensity is 2500-3000 Lx, and the illumination is 8-14 hours for light-dark alternate culture.
3. The method of claim 1, wherein the primary induction medium is formulated as follows: WPM+NAA 0.04 mg/L+ZT1.1 mg/L+6-BA 3.5mg/L+GA 3 0.15mg/L+ Activity1.0g/L of charcoal.
4. The method of claim 1, wherein the formulation of the secondary proliferation medium is: WPM+NAA 0.01mg/L+TDZ 0.9mg/L+ZT 0.8 mg/L+urea 0.08g/L.
5. The method of claim 1, wherein the rooting medium is formulated as follows: SH+IBA 0.3mg/L+IAA 1.2mg/L+PPP 333 0.04 mg/L+150 mg/L peptone+0.2 g/L activated carbon.
6. Use of the method according to any one of claims 1-5 in the tissue culture and rapid propagation of phyllanthus emblica seedlings.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202410278531.0A CN117859652A (en) | 2024-03-12 | 2024-03-12 | Fructus phyllanthi tissue culture rapid propagation seedling method and application |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202410278531.0A CN117859652A (en) | 2024-03-12 | 2024-03-12 | Fructus phyllanthi tissue culture rapid propagation seedling method and application |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN117859652A true CN117859652A (en) | 2024-04-12 |
Family
ID=90584877
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202410278531.0A Pending CN117859652A (en) | 2024-03-12 | 2024-03-12 | Fructus phyllanthi tissue culture rapid propagation seedling method and application |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN117859652A (en) |
Citations (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2000057690A2 (en) * | 1999-03-25 | 2000-10-05 | University Of Guelph | Micropropagation and production of phytopharmaceutical plants |
| US6667065B1 (en) * | 1998-04-01 | 2003-12-23 | Danisco A/S | Non-maltogenic exoamylases and their use in retarding retrogradation of starch |
| CN1843097A (en) * | 2006-03-13 | 2006-10-11 | 西北大学 | High effective propagation of blattbulume stem node and knottiness axillary bud tissue culture seedling |
| CN106266391A (en) * | 2016-09-05 | 2017-01-04 | 佛山市顺德区宝铜金属科技有限公司 | A kind of medicine for fat-eliminating slimming |
| CN107960328A (en) * | 2017-12-21 | 2018-04-27 | 中国林业科学研究院资源昆虫研究所 | A kind of method of emblic tissue cultures |
| CN108040773A (en) * | 2017-12-13 | 2018-05-18 | 福建省农业科学院果树研究所 | A kind of method for cultivating emblic seedling |
| CN211210904U (en) * | 2019-11-12 | 2020-08-11 | 云南省农业科学院热区生态农业研究所 | Phyllanthus emblica tissue culture integrated device |
-
2024
- 2024-03-12 CN CN202410278531.0A patent/CN117859652A/en active Pending
Patent Citations (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US6667065B1 (en) * | 1998-04-01 | 2003-12-23 | Danisco A/S | Non-maltogenic exoamylases and their use in retarding retrogradation of starch |
| WO2000057690A2 (en) * | 1999-03-25 | 2000-10-05 | University Of Guelph | Micropropagation and production of phytopharmaceutical plants |
| CN1843097A (en) * | 2006-03-13 | 2006-10-11 | 西北大学 | High effective propagation of blattbulume stem node and knottiness axillary bud tissue culture seedling |
| CN106266391A (en) * | 2016-09-05 | 2017-01-04 | 佛山市顺德区宝铜金属科技有限公司 | A kind of medicine for fat-eliminating slimming |
| CN108040773A (en) * | 2017-12-13 | 2018-05-18 | 福建省农业科学院果树研究所 | A kind of method for cultivating emblic seedling |
| CN107960328A (en) * | 2017-12-21 | 2018-04-27 | 中国林业科学研究院资源昆虫研究所 | A kind of method of emblic tissue cultures |
| CN211210904U (en) * | 2019-11-12 | 2020-08-11 | 云南省农业科学院热区生态农业研究所 | Phyllanthus emblica tissue culture integrated device |
Non-Patent Citations (2)
| Title |
|---|
| VERMA, BHANU等: "Micropropagation of Emblica officinalis Gaertn. through mature nodal explants", JOURNAL OF PHYTOLOGICAL RESEARCH, vol. 9, no. 2, 31 December 1996 (1996-12-31), pages 107 * |
| 张守英, 姚小华, 任华东, 王开良: "余甘子丛生芽诱导和快速繁殖研究", 经济林研究, no. 01, 30 March 2002 (2002-03-30), pages 12 * |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN108522277B (en) | Tissue culture seedling raising method for semi-lignified stem segments with buds of 'Hongyang' kiwi fruits | |
| CN110810250B (en) | Method for rapid propagation in tissue culture of date palm | |
| CN105794642A (en) | Method for efficiently and rapidly regenerating adventitious buds from pear leaves | |
| CN113080063B (en) | Rapid rooting method for tissue culture of coarse chaff tree | |
| CN114532226A (en) | Culture medium combination and method for tissue culture and rapid propagation of curculigo orchioides | |
| CN107135945B (en) | Tissue culture medium of linden tree and rapid propagation method thereof | |
| CN109729976B (en) | Tissue culture and rapid propagation method for phoenix-tail bamboos | |
| CN117859652A (en) | Fructus phyllanthi tissue culture rapid propagation seedling method and application | |
| CN115968786A (en) | Culture medium and culture method for tea tree tissue culture | |
| CN108739403B (en) | Tissue culture rapid propagation method of rosewood | |
| KR100865680B1 (en) | The mass producing method of regenerated plant from the leaf segment of ficus carica | |
| CN114698549B (en) | Tissue culture medium and tissue culture method for rapid propagation of grape stock stem segments | |
| CN113383706B (en) | Efficient eucommia bark regeneration method based on LED light quality regulation | |
| CN115885852A (en) | Tissue culture method for leaf of chicory | |
| CN115152629A (en) | Raspberry tissue culture method | |
| CN107853179A (en) | A kind of tissue culture and rapid propagation method of orchid | |
| CN111771725A (en) | Optimized ginger virus-free seedling regeneration propagation method and development of industrialization process thereof | |
| CN113100065A (en) | Induction medium for tissue culture and rapid propagation of dracocephalum plants and application thereof | |
| CN115836646B (en) | Tissue culture method of idesia polycarpa | |
| CN115517167A (en) | Culture method for inducing blueberry chromosome doubling | |
| CN116138168B (en) | Culture medium for promoting germination of raspberry seeds and tissue culture seedling method | |
| CN111387057B (en) | Gymnadenia conopsea sterile seeding culture medium | |
| CN117322329B (en) | Culture method of bulbil konjak | |
| CN114097619B (en) | In-vitro conservation method for sugarcane germplasm resources | |
| CN117099686B (en) | North American ilex tissue culture rapid propagation method and application |
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 |