CN105684898A - Method for efficiently inducing hybrid sandalwood body cell embryo to generate and regenerate plant - Google Patents
Method for efficiently inducing hybrid sandalwood body cell embryo to generate and regenerate plant Download PDFInfo
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- CN105684898A CN105684898A CN201610038247.1A CN201610038247A CN105684898A CN 105684898 A CN105684898 A CN 105684898A CN 201610038247 A CN201610038247 A CN 201610038247A CN 105684898 A CN105684898 A CN 105684898A
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- 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
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- 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
- A01H4/001—Culture apparatus for tissue culture
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Abstract
The invention discloses a method for efficiently inducing a hybrid sandalwood body cell embryo to generate and regenerate a plant. The method is characterized in that an excellent cultivation hybrid sandalwood tender stem is selected as an explant; graded disinfecting procedures are adopted for surface disinfection of the explant, so that a higher success rate can be obtained; the successfully-disinfected aseptic explant is cultured on an embryonic callus culture medium containing 2,4-D to induce a fibrous embryonic callus; the fibrous embryonic callus is transferred to a somatic embryo for inducing, maturing and germinating as well as culturing on a plant regeneration culture medium; a regenerated plant can be obtained after three months. The method has the characteristics of short reproduction cycle, high quantity and high stability; the problem that the propagation efficiency of seedlings cultured and propagated by hybrid sandalwood tissues is low is solved, a seedling raising technique is provided for developing rapid propagation of hybrid sandalwood seedlings, and a regeneration path is also provided for high-efficiency gene transformation. The method disclosed by the invention has the advantages of high operability, high propagation efficiency, high application value and the like.
Description
Technical field:
The invention belongs to field of plant reproduction, be specifically related to a kind of efficient induction hybrid Lignum Santali Albi somatic embryo and the method with plant regeneration occurs。
Background technology:
Lignum Santali Albi (SantalumalbumL.) belongs to (Santalum) semiparasite aiphyllium for Santalaceae (Santalaceae) Lignum Santali Albi, for famous and precious, rare plant, it is distributed mainly on India, Indonesia, Australian and more Pacific archipelagos。Its history in existing thousands of years of human use, especially famous because its timber can be used for carving and refine Oleum Santali albi, have the title of " green gold "。The Chinese primary distribution without Lignum Santali Albi, its finished product or semi-finished product are in the past always from external import。Within 1962, South China Botanical Garden Chinese Academy of Sciences introduces Lignum Santali Albi seed growing success from Indonesia first in continent, introduces " Laoshan is fragrant " high-quality kind source from India thereafter and also succeeds。Through the development test of decades, nowadays Lignum Santali Albi can have been set seeds in South China Botanical Garden and bred and energy Edgeworthia chrysantha Lindl.。Recent two decades comes, and our seminar also promotes to other area domestic and plants experimentally Lignum Santali Albi。
Research data shows: in Fiji, and the sexual hybridization kind F1 generation (hybrid Lignum Santali Albi S.album × S.yasi) of Lignum Santali Albi and Fiji Lignum Santali Albi shows obvious hybrid vigor, and its growth potential is higher by 300% than maternal Fiji Lignum Santali Albi, then higher by 32% than male parent Lignum Santali Albi；Edgeworthia chrysantha Lindl. aspect, Lignum Santali Albi needs 10 years, and hybrid Lignum Santali Albi then needs 6-7；In heartwood growth, general every strain Lignum Santali Albi needs plantation 25-30 heartwood to reach 40-80 kilogram, and hybrid Lignum Santali Albi F1 only need to plant 15-20。We strengthen and on the basis exchanged both at home and abroad, want Lignum Santali Albi study base, Dendrocalamus giganteus Munro island research worker from overseas introduction hybrid Lignum Santali Albi to Lignum Santali Albi study base, Dendrocalamus giganteus Munro island, well-grown with Zhaoqing Guangdong height in recent years；Although hybrid Lignum Santali Albi can be solid on a small quantity, but characters of progenies seriously separates, it is necessary to seek pierre technology to maintain hybrid vigor。
At present, have no hybrid Lignum Santali Albi somatic embryo both at home and abroad to occur and plant regeneration report and patent application。
Summary of the invention:
It is an object of the invention to provide a kind of strong operability, reproductive efficiency and using value high, can provide the efficiently induction hybrid Lignum Santali Albi somatic embryo of seedling that the method with plant regeneration occurs for the extensive Fast-propagation of hybrid Lignum Santali Albi
The present invention adopts hybrid Lignum Santali Albi tender stem to be outer implant, carries out somatic embryo induction, sprouting, maturation, regeneration plant, can obtain quantity by embryo callus subculture somatic embryos many, and breeding is fast, stability high。For successfully carrying out hybrid Lignum Santali Albi seedling Fast-propagation offer seedling-raising technique and a kind of effective path for transformation can be provided for transgenic experiments, it is achieved thereby that the purpose of the present invention。
There is the method with plant regeneration in the efficiently induction hybrid Lignum Santali Albi somatic embryo of the present invention, it is characterised in that comprises the following steps:
A, the induction of embryo callus and propagation: using the young tender stem of hybrid Lignum Santali Albi as outer implant, carry out disinfection, outer implant after sterilization is seeded in embryonic callus induction culture medium and cultivates, obtain embryo callus, embryo callus is transferred in new embryonic callus induction culture medium and carries out subculture multiplication cultivation, obtain cells,primordial, every 2 weeks transfer 1 time, the regeneration capacity of embryo callus subculture can be maintained, described embryonic callus induction culture medium every liter contains 2, 4-D (2, 4-dichlorphenoxyacetic acid) 0.5-1.0mg, sucrose 20-30g, agar 6-7g, surplus is MS culture medium, described hybrid Lignum Santali Albi is the sexual hybridization kind F1 generation of male parent Lignum Santali Albi (Santalumalbum) and female parent Fiji Lignum Santali Albi (Santalumyasi)；
B, the induction of somatic embryo is with ripe: cells,primordial is transferred on somatic embryo inducing culture the generation of somatic embryos, obtain somatic embryo, select somatic embryo and proceed to cultivation in somatic embryo maturation culture medium, obtain ripe cotyledonary embryos, described somatic embryo inducing culture every liter contains 6-BA (6-benzyl aminoadenine) 0.2-0.5mg, sucrose 20-30g, agar 6-7g, surplus is MS culture medium, described somatic embryo maturation culture medium every liter contains 6-BA (6-benzyl aminoadenine) 0.05-0.1mg, sucrose 20-30g, agar 6-7g, surplus is MS culture medium；
C, somatic embryo are sprouted and plant regeneration: the cotyledonary embryos of picking maturation proceeds in somatic embryo germination medium to be cultivated, obtain the somatic embryo sprouted, the somatic embryo of sprouting is transferred to plant regeneration culture medium is cultivated, obtaining complete regeneration plant, described somatic embryo germination medium every liter contains 6-BA (6-benzyl aminoadenine) 0.2-0.5mg, GA3(gibberellins) 0.5-1mg, sucrose 20-30g, agar 6-7g, surplus is MS culture medium, described plant regeneration culture medium every liter contains 6-BA (6-benzyl aminoadenine) 0.2-1mg, IBA (indolebutyric acid) 0.2-1mg, sucrose 20-30g, agar 6-7g, and surplus is 1/2MS culture medium。
Described using the young tender stem of hybrid Lignum Santali Albi as outer implant, carry out disinfection, preferably by outer implant first with implant 2-3 time outside the alcohol water blend wiping of volume fraction 50%, then first soak 30 seconds with volume fraction 75% alcohol water blend on superclean bench, immersion 5～6min in mass fraction 0.1%HgC1 solution is put into after aseptic water washing, 1 time is rocked every 2 minutes, aseptic water washing 2-3 time, again with volume fraction 6-12% hydrogenperoxide steam generator sterilization 6-8min, aseptic water washing 3-5 time, it is thus achieved that the outer implant after sterilization。
Preferably, the outer implant after sterilization is seeded in embryonic callus induction culture medium and cultivates by described step a, it is thus achieved that embryo callus, is transferred to by embryo callus in new embryonic callus induction culture medium and carries out subculture multiplication cultivation；
Cells,primordial is transferred on somatic embryo inducing culture the generation of somatic embryos by described step b, it is thus achieved that somatic embryo, selects somatic embryo and proceeds in somatic embryo maturation culture medium and cultivate；
The cotyledonary embryos of the picking maturation of described step c proceeds in somatic embryo germination medium to be cultivated, it is thus achieved that the somatic embryo of sprouting, is transferred to by the somatic embryo of sprouting and cultivates in plant regeneration culture medium；
Its condition of culture is all intensity of illumination 1000-2000lx, light application time 10h/d, cultivation temperature 26 ± 2 DEG C。
MS culture medium is international culture medium, its composition and collocation method are shown in MurashigeT, SkoogF (1962) (Arevisedmediumforrapidgrowthandbioassaywithtobaccotissue cultures.PhysiolPlant15:473 497)。1/2MS culture medium refers to that by a great number of elements consumption in MS culture medium be original 1/2, and all the other compositions are constant。
The present invention selects excellent cultivation hybrid Lignum Santali Albi children's tender stem as outer implant, adopts the surface sterilization that gradation disinfectant program carries out outer implant can obtain higher success rate。Sterilize successful aseptic explant containing 2, the embryo callus culture medium of 4-D is cultivated, induce threadiness embryo callus, it is then transferred in somatic embryo inducement, maturation, sprouting, plant regeneration culture medium and cultivates, regeneration plant can be obtained after 3 months, breeding cycle is short, quantity is many, stability high feature, the reproductive efficiency solving to exist in hybrid Lignum Santali Albi tissue culture propagating seedling is not high, will for carrying out hybrid Lignum Santali Albi seedling Fast-propagation offer seedling-raising technique, it is possible to provide Regeneration Ways for efficient gene transformation。The present invention has strong operability, reproductive efficiency and using value advantages of higher。
Accompanying drawing illustrates:
Fig. 1 be hybrid Lignum Santali Albi somatic embryo occur and plant regeneration, wherein A, embryo callus；B, induction somatic embryo；Each stage that C, somatic embryo are grown: globular embryo, heart-shape embryo, torpedo-shape embryo, cotyledon shape embryo, ripe cotyledonary embryos；D regeneration plant。A-C scale=2 millimeter, D scale=1 centimetre。
Detailed description of the invention:
Following example are further illustrating the present invention, rather than limitation of the present invention。
Embodiment 1: efficiently the method with plant regeneration occurs induction hybrid Lignum Santali Albi somatic embryo, comprises the following steps:
(1) outer implant and process thereof
Pick up from Guangdong height and take Sandalwood cultivation base, the Dendrocalamus giganteus Munro island introducing and planting elite plant strain hybrid Lignum Santali Albi of 6 years (the sexual hybridization kind F1 generation of male parent Lignum Santali Albi (Santalumalbum) and female parent Fiji Lignum Santali Albi (Santalumyasi)) tender stem section as outer implant, first use volume fraction 50% alcohol water blend wiping outer implant 2-3 time, then first soak 30 seconds with volume fraction 75% alcohol water blend on superclean bench, immersion about 5-6min in mass fraction 0.1%HgC1 solution is put into after aseptic water washing, 1 time is rocked every 2 minutes, aseptic water washing 2-3 time, sterilize about 6-8min with volume fraction 6% hydrogenperoxide steam generator again, aseptic water washing 3-5 time, obtain the tender stem after surface sterilization processes。
(2) induction of embryo callus and propagation
Tender stem after surface sterilization being processed with sharp sterile scalpel is cut into the band knot stem section of 1-1.5cm, and aseptic nipper places stem section on embryonic callus induction culture medium。Condition of culture, intensity of illumination 1000lx, light application time 10h/d, cultivation temperature 26 ± 2 DEG C。After 7-10d, stem section starts to expand and incision starts to sprout, and cultivates the embryo callus (Fig. 1-A) that visible division is vigorous after 20d, and callus induction rate is up to more than 80%。Then embryo callus is transferred in new embryonic callus induction culture medium and carries out subculture multiplication cultivation, it is thus achieved that cells,primordial。Every 2 weeks transfer 1 time, can maintain the regeneration capacity of embryo callus subculture。Described embryonic callus induction culture medium every liter contains 2,4-D (2,4-dichlorphenoxyacetic acid) 1.0mg, sucrose 30g, agar 7g, and surplus is MS culture medium, pH5.8, and its compound method is after composition mix homogeneously, will to adjust pH value, and sterilizing is standby。
(3) induction of somatic embryo is with ripe
Take the cells,primordial of 6d, the generation of somatic embryos on somatic embryo inducing culture after successive transfer culture。Condition of culture, intensity of illumination 1000lx, light application time 10h/d, cultivation temperature 26 ± 2 DEG C。The appearance of the visible somatic embryo of naked eyes after 15d, during to 30d, it is seen that to substantial amounts of somatic embryo (Fig. 1-B)。Select somatic embryo and proceed to somatic embryo maturation culture medium, condition of culture, intensity of illumination 1000lx, light application time 10h/d, cultivation temperature 26 ± 2 DEG C。Obtain ripe cotyledonary embryos (its growth course is such as shown in Fig. 1-C)。
Described somatic embryo inducing culture every liter contains 6-BA (6-benzyl aminoadenine) 0.2mg, sucrose 30g, agar 7g, and surplus is MS culture medium, pH5.8, and its compound method is after composition mix homogeneously, will to adjust pH value, and sterilizing is standby。
Described somatic embryo maturation culture medium every liter contains 6-BA (6-benzyl aminoadenine) 0.1mg, sucrose 30g, agar 6g, and surplus is MS culture medium, pH5.8, and its compound method is after composition mix homogeneously, will to adjust pH value, and sterilizing is standby。
(4) somatic embryo is sprouted and plant regeneration
The ripe cotyledonary embryos selecting about 1-2cm proceeds to cultivation, intensity of illumination 1000lx, light application time 10h/d on somatic embryo germination medium, cultivation temperature 26 ± 2 DEG C, cultivate about 2 weeks two panels cotyledons to start to expand, and start to grow the root that children is tender, it is thus achieved that the somatic embryo after sprouting。Somatic embryo after sprouting is transferred in plant regeneration culture medium, intensity of illumination 1000lx, light application time 10h/d, cultivation temperature 26 ± 2 DEG C, cultivates 2 weeks, buds into complete regeneration plant (Fig. 1-D) further。
Described somatic embryo germination medium every liter contains 6-BA (6-benzyl aminoadenine) 0.5mg, GA3(gibberellins) 1mg, sucrose 30g, agar 7g, surplus is MS culture medium, pH5.8, and its compound method is after composition mix homogeneously, will to adjust pH value, and sterilizing is standby。
Described plant regeneration culture medium every liter contains 6-BA (6-benzyl aminoadenine) 1mg, IBA (indolebutyric acid) 0.2mg, sucrose 30g, agar 7g, surplus is 1/2MS culture medium, pH5.8, and its compound method is by after composition mix homogeneously, adjusting pH value, sterilizing is standby。
Embodiment 2: efficiently the method with plant regeneration occurs induction hybrid Lignum Santali Albi somatic embryo, comprises the following steps:
(1) outer implant and process thereof
Pick up from Guangdong height and take Sandalwood cultivation base, the Dendrocalamus giganteus Munro island introducing and planting elite plant strain hybrid Lignum Santali Albi of 6 years (the sexual hybridization kind F1 generation of male parent Lignum Santali Albi (Santalumalbum) and female parent Fiji Lignum Santali Albi (Santalumyasi)) tender stem section as outer implant, first use volume fraction 50% alcohol water blend wiping outer implant 2-3 time, then first soak 30 seconds with volume fraction 75% alcohol water blend on superclean bench, immersion about 5-6min in mass fraction 0.1%HgC1 solution is put into after aseptic water washing, 1 time is rocked every 2 minutes, aseptic water washing 2-3 time, sterilize about 6-8min with volume fraction 12% hydrogenperoxide steam generator again, aseptic water washing 3-5 time, obtain the tender stem after surface sterilization processes。
(2) induction of embryo callus and propagation
Tender stem after surface sterilization being processed with sharp sterile scalpel is cut into the band knot stem section of 1-1.5cm, and aseptic nipper places stem section on embryonic callus induction culture medium。Condition of culture, intensity of illumination 2000lx, light application time 10h/d, cultivation temperature 26 ± 2 DEG C。After 7-10d, stem section starts to expand and incision starts to sprout, and cultivates the embryo callus that visible division is vigorous after 20d, and callus induction rate is up to more than 80%。Then embryo callus is transferred in new embryonic callus induction culture medium and carries out subculture multiplication cultivation, it is thus achieved that cells,primordial。Every 2 weeks transfer 1 time, can maintain the regeneration capacity of embryo callus subculture。Described embryonic callus induction culture medium every liter contains 2,4-D (2,4-dichlorphenoxyacetic acid) 0.5mg, sucrose 20g, agar 6g, and surplus is MS culture medium, pH5.8, and its compound method is after composition mix homogeneously, will to adjust pH value, and sterilizing is standby。
(3) induction of somatic embryo is with ripe
Take the cells,primordial of 6d, the generation of somatic embryos on somatic embryo inducing culture after successive transfer culture。Condition of culture, intensity of illumination 2000lx, light application time 10h/d, cultivation temperature 26 ± 2 DEG C。The appearance of the visible somatic embryo of naked eyes after 15d, during to 30d, it is seen that to substantial amounts of somatic embryo。Select somatic embryo and proceed to somatic embryo maturation culture medium, condition of culture, intensity of illumination 2000lx, light application time 10h/d, cultivation temperature 26 ± 2 DEG C。Obtain ripe cotyledonary embryos。
Described somatic embryo inducing culture every liter contains 6-BA (6-benzyl aminoadenine) 0.5mg, sucrose 20g, agar 6g, and surplus is MS culture medium, pH5.8, and its compound method is after composition mix homogeneously, will to adjust pH value, and sterilizing is standby。
Described somatic embryo maturation culture medium every liter contains 6-BA (6-benzyl aminoadenine) 0.05mg, sucrose 20g, agar 7g, and surplus is MS culture medium, pH5.8, and its compound method is after composition mix homogeneously, will to adjust pH value, and sterilizing is standby。
(4) somatic embryo is sprouted and plant regeneration
The ripe cotyledonary embryos selecting about 1-2cm proceeds to cultivation, intensity of illumination 2000lx, light application time 10h/d on somatic embryo germination medium, cultivation temperature 26 ± 2 DEG C, cultivate about 2 weeks two panels cotyledons to start to expand, and start to grow the root that children is tender, it is thus achieved that the somatic embryo after sprouting。Somatic embryo after sprouting is transferred in plant regeneration culture medium, intensity of illumination 2000lx, light application time 10h/d, cultivation temperature 26 ± 2 DEG C, cultivates 2 weeks, buds into complete regeneration plant further。
Described somatic embryo germination medium every liter contains 6-BA (6-benzyl aminoadenine) 0.2mg, GA3(gibberellins) 0.5mg, sucrose 20g, agar 6g, surplus is MS culture medium, pH5.8, and its compound method is after composition mix homogeneously, will to adjust pH value, and sterilizing is standby。
Described plant regeneration culture medium every liter contains 6-BA (6-benzyl aminoadenine) 0.2mg, IBA (indolebutyric acid) 1mg, sucrose 20g, agar 6g, surplus is 1/2MS culture medium, pH5.8, and its compound method is by after composition mix homogeneously, adjusting pH value, sterilizing is standby。
Claims (3)
1. there is the method with plant regeneration in an efficient induction hybrid Lignum Santali Albi somatic embryo, it is characterised in that comprises the following steps:
A, the induction of embryo callus and propagation: using the young tender stem of hybrid Lignum Santali Albi as outer implant, carry out disinfection, outer implant after sterilization is seeded in embryonic callus induction culture medium and cultivates, obtain embryo callus, embryo callus is transferred in new embryonic callus induction culture medium and carries out subculture multiplication cultivation, obtain cells,primordial, described embryonic callus induction culture medium every liter contains 2, 4-D0.5-1.0mg, sucrose 20-30g, agar 6-7g, surplus is MS culture medium, described hybrid Lignum Santali Albi is the sexual hybridization kind F1 generation of male parent Lignum Santali Albi (Santalumalbum) and female parent Fiji Lignum Santali Albi (Santalumyasi)；
B, somatic embryo induction with ripe: cells,primordial is transferred on somatic embryo inducing culture the generation of somatic embryos, obtain somatic embryo, select somatic embryo and proceed to cultivation in somatic embryo maturation culture medium, obtain ripe cotyledonary embryos, described somatic embryo inducing culture every liter contains 6-BA0.2-0.5mg, sucrose 20-30g, agar 6-7g, surplus is MS culture medium, described somatic embryo maturation culture medium every liter contains 6-BA0.05-0.1mg, sucrose 20-30g, agar 6-7g, and surplus is MS culture medium；
C, somatic embryo are sprouted and plant regeneration: the cotyledonary embryos of picking maturation proceeds in somatic embryo germination medium to be cultivated, obtain the somatic embryo sprouted, the somatic embryo of sprouting is transferred to plant regeneration culture medium is cultivated, obtaining complete regeneration plant, described somatic embryo germination medium every liter contains 6-BA0.2-0.5mg, GA30.5-1mg, sucrose 20-30g, agar 6-7g, surplus is MS culture medium, and described plant regeneration culture medium every liter contains 6-BA0.2-1mg, IBA0.2-1mg, sucrose 20-30g, agar 6-7g, and surplus is 1/2MS culture medium。
2. there is the method with plant regeneration in efficient induction hybrid Lignum Santali Albi somatic embryo according to claim 1, it is characterized in that, described using the young tender stem of hybrid Lignum Santali Albi as outer implant, carry out disinfection, it is first with implant 2-3 time outside the alcohol water blend wiping of volume fraction 50% by outer implant, then first soak 30 seconds with volume fraction 75% alcohol water blend on superclean bench, immersion 5～6min in mass fraction 0.1%HgC1 solution is put into after aseptic water washing, 1 time is rocked every 2 minutes, aseptic water washing 2-3 time, again with volume fraction 6-12% hydrogenperoxide steam generator sterilization 6-8min, aseptic water washing 3-5 time, obtain the outer implant after sterilization。
3. there is the method with plant regeneration in efficient induction hybrid Lignum Santali Albi somatic embryo according to claim 1, it is characterised in that
Outer implant after sterilization is seeded in embryonic callus induction culture medium and cultivates by described step a, it is thus achieved that embryo callus, is transferred to by embryo callus in new embryonic callus induction culture medium and carries out subculture multiplication cultivation；
Cells,primordial is transferred on somatic embryo inducing culture the generation of somatic embryos by described step b, it is thus achieved that somatic embryo, selects somatic embryo and proceeds in somatic embryo maturation culture medium and cultivate；
The cotyledonary embryos of the picking maturation of described step c proceeds in somatic embryo germination medium to be cultivated, it is thus achieved that the somatic embryo of sprouting, is transferred to by the somatic embryo of sprouting and cultivates in plant regeneration culture medium；
Its condition of culture is all intensity of illumination 1000-2000lx, light application time 10h/d, cultivation temperature 26 ± 2 DEG C。
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN108207385A (en) * | 2018-02-09 | 2018-06-29 | 中国林业科学研究院热带林业研究所 | A kind of chemical regulation method for improving santal seedling quality and resistance |
| CN112586349A (en) * | 2020-11-11 | 2021-04-02 | 江西省中国科学院庐山植物园 | Method for rapidly propagating clematis chinensis seedlings through somatic embryogenesis |
| CN115250913A (en) * | 2022-07-25 | 2022-11-01 | 三峡大学 | Rhus chinensis somatic embryogenesis and plant regeneration method |
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2016
- 2016-01-20 CN CN201610038247.1A patent/CN105684898B/en not_active Expired - Fee Related
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108207385A (en) * | 2018-02-09 | 2018-06-29 | 中国林业科学研究院热带林业研究所 | A kind of chemical regulation method for improving santal seedling quality and resistance |
| CN112586349A (en) * | 2020-11-11 | 2021-04-02 | 江西省中国科学院庐山植物园 | Method for rapidly propagating clematis chinensis seedlings through somatic embryogenesis |
| CN115250913A (en) * | 2022-07-25 | 2022-11-01 | 三峡大学 | Rhus chinensis somatic embryogenesis and plant regeneration method |
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