CN109197598A - A kind of Populus Tomentosa culture method - Google Patents
A kind of Populus Tomentosa culture method Download PDFInfo
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- CN109197598A CN109197598A CN201811453977.3A CN201811453977A CN109197598A CN 109197598 A CN109197598 A CN 109197598A CN 201811453977 A CN201811453977 A CN 201811453977A CN 109197598 A CN109197598 A CN 109197598A
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- 241000249899 Populus tomentosa Species 0.000 title claims abstract description 47
- 238000012136 culture method Methods 0.000 title claims abstract description 19
- 230000006698 induction Effects 0.000 claims abstract description 150
- 239000001963 growth medium Substances 0.000 claims abstract description 102
- 230000002062 proliferating effect Effects 0.000 claims abstract description 57
- 230000035755 proliferation Effects 0.000 claims abstract description 22
- 229920001817 Agar Polymers 0.000 claims description 23
- 239000008272 agar Substances 0.000 claims description 23
- 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 22
- 229930006000 Sucrose Natural products 0.000 claims description 22
- 239000005720 sucrose Substances 0.000 claims description 22
- 230000004069 differentiation Effects 0.000 claims description 21
- 239000002609 medium Substances 0.000 claims description 21
- 238000005286 illumination Methods 0.000 claims description 15
- 241000196324 Embryophyta Species 0.000 claims description 12
- 206010020649 Hyperkeratosis Diseases 0.000 claims description 8
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 8
- 239000007921 spray Substances 0.000 claims description 7
- 238000012549 training Methods 0.000 claims description 5
- 230000008901 benefit Effects 0.000 claims description 4
- 238000000034 method Methods 0.000 abstract description 15
- 239000000463 material Substances 0.000 abstract description 5
- 238000004161 plant tissue culture Methods 0.000 abstract description 5
- 238000004659 sterilization and disinfection Methods 0.000 abstract description 4
- 238000004643 material aging Methods 0.000 abstract description 3
- 239000005556 hormone Substances 0.000 abstract description 2
- 229940088597 hormone Drugs 0.000 abstract description 2
- 230000001954 sterilising effect Effects 0.000 abstract description 2
- 230000000763 evoking effect Effects 0.000 abstract 1
- 238000009395 breeding Methods 0.000 description 10
- 230000001488 breeding effect Effects 0.000 description 10
- 241000219000 Populus Species 0.000 description 9
- 210000001519 tissue Anatomy 0.000 description 9
- 230000001939 inductive effect Effects 0.000 description 4
- 238000004519 manufacturing process Methods 0.000 description 4
- 240000000111 Saccharum officinarum Species 0.000 description 3
- 235000007201 Saccharum officinarum Nutrition 0.000 description 3
- 238000005516 engineering process Methods 0.000 description 3
- 230000006872 improvement Effects 0.000 description 3
- 241000168036 Populus alba Species 0.000 description 2
- 210000004027 cell Anatomy 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 230000004083 survival effect Effects 0.000 description 2
- 244000050510 Cunninghamia lanceolata Species 0.000 description 1
- 241001465754 Metazoa Species 0.000 description 1
- 240000002834 Paulownia tomentosa Species 0.000 description 1
- 230000002159 abnormal effect Effects 0.000 description 1
- 230000032683 aging Effects 0.000 description 1
- 230000011681 asexual reproduction Effects 0.000 description 1
- 238000013465 asexual reproduction Methods 0.000 description 1
- 238000010924 continuous production Methods 0.000 description 1
- 238000001784 detoxification Methods 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 230000007613 environmental effect Effects 0.000 description 1
- 239000010977 jade Substances 0.000 description 1
- 210000000056 organ Anatomy 0.000 description 1
- 238000003976 plant breeding Methods 0.000 description 1
- 230000008121 plant development Effects 0.000 description 1
- 238000004321 preservation Methods 0.000 description 1
- 230000008569 process Effects 0.000 description 1
- 230000001902 propagating effect Effects 0.000 description 1
- 210000001938 protoplast Anatomy 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 230000000284 resting effect Effects 0.000 description 1
- 229930000044 secondary metabolite Natural products 0.000 description 1
- 241000894007 species Species 0.000 description 1
- 238000004017 vitrification Methods 0.000 description 1
Classifications
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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
-
- 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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- Life Sciences & Earth Sciences (AREA)
- Developmental Biology & Embryology (AREA)
- Engineering & Computer Science (AREA)
- Biotechnology (AREA)
- Cell Biology (AREA)
- Botany (AREA)
- Environmental Sciences (AREA)
- Breeding Of Plants And Reproduction By Means Of Culturing (AREA)
Abstract
The present invention provides a kind of Populus Tomentosa culture methods, are related to field of plant tissue culture technique.The method of the invention is inoculated in progress proliferative induction culture on proliferative induction culture medium the following steps are included: be woven to explant with Populus Tomentosa, by the explant, obtains proliferation bud；The proliferation bud is inoculated in progress root induction culture on root induction culture medium, obtains Chinese white poplar tissue-cultured seedling.The method of the invention does not include the Initial culture of traditional sense, but the S1 or S2 that explant material is directly directly inoculated according to material aging degree to addition hormone after sterilization are cultivated, evoking adventive bud generates proliferation bud, to carry out culture of rootage, scheme is simple, tissue culture is high-efficient, and explant breaks up bud induction rate up to 90%, and rooting induction rate is up to 88.5%.
Description
Technical field
The invention belongs to field of plant tissue culture technique, and in particular to a kind of tissue culture method of Chinese white poplar.
Background technique
Plant tissue culture technique is based on the totipotency of cell, refer to aseptically, by vitro plant organ,
Tissue, cell and protoplast are cultivated and give suitable environmental condition on artificial medium, induction generation callus,
Resting bud etc., and then cultivate the technology at intact plant.In recent years, plant tissue culture technique becomes plant breeding, plant
The core technology of detoxification and the research of fast breeding, Secondary metabolites production, Preservation of plant germplasin etc., training
The new variety of plant of large area plantation is brought out.Currently, plant tissue culture technique agricultural, forestry, animal husbandry, etc. it is multiple
Field, which has, to be widely applied, and group culturation rapid propagating technology can not be limited by conditions such as seasons, breeding speed short with growth cycle
Degree is fast and nursery stock neat and consistent, can whole year production the advantages that.Such as in terms of forestry, paulownia, Chinese white poplar, China fir etc. have put into work
Factoryization is fast numerous, greatly improves the improvement of yield and quality, so that tissue-cultured seedling production is formed industrialization in domestic and international market, obtains
Splendid economic benefit is taken.
Chinese white poplar (Populus tomentosa) is Salicaceae Populus fallen leaves megaphanerophyte, and trunk straight elevator is thick with leaves, fits
Ying Xingqiang is the important quick growing species of trees of northern China, is widely used in industrial cut stock and urban afforestation.However the hair in nursery stock production
White poplar is usually bred with asexual reproduction methods such as cuttage, press strips, is difficult to take root, and propagation material is limited in addition, not only consumptive material
It is more, time-consuming, and survival rate is low, and breeding coefficient is small, and breeding a large amount of nursery stocks in a short time can be very difficult.
Summary of the invention
In view of this, being suitable for different genotype, difference the purpose of the present invention is to provide a kind of Populus Tomentosa culture method
The Chinese white poplar explant of maturity can obtain a large amount of Chinese white poplar nursery stocks, high survival rate, breeding coefficient height in a short time.
In order to achieve the above-mentioned object of the invention, the present invention the following technical schemes are provided:
The present invention provides a kind of Populus Tomentosa culture methods, comprising the following steps:
1) explant is woven to Populus Tomentosa, the explant is inoculated on proliferative induction culture medium and carries out proliferative induction
Culture, obtains proliferation bud；The proliferative induction culture using proliferative induction culture medium S1 carry out, the proliferative induction culture medium S1 with
MS culture medium is minimal medium, further includes: the sucrose of the IBA of the 6-BA of 0.2mg/L, 0.1mg/L, 24~35g/L and 4.5~
The agar of 8.6g/L；The temperature of the proliferative induction culture is (22 ± 5) DEG C, and intensity of illumination is 1000~5000lx, when illumination
Between be 12h/d；
2) the proliferation bud is inoculated in progress root induction culture on root induction culture medium, obtains Chinese white poplar tissue-cultured seedling；
The root induction culture is carried out using root induction culture medium R1, and the root induction culture medium R1 is with 1/2MS culture medium
Minimal medium further includes the IBA of 0.3mg/L, the sucrose of 10~20g/L and the agar of 4.5~8.6g/L；The root induction
The temperature of culture is (22 ± 5) DEG C, and intensity of illumination is 1000~5000lx, light application time 12h/d.
Preferably, the explant includes annotinous branch or the spray that will grow after diameter≤3cm bough water planting.
Preferably, when the explant is the spray obtained with ancient tree bough water planting, with proliferative induction culture medium S1
It further include carrying out Bud Differentiation culture using proliferative induction culture medium S2 before carrying out proliferative induction culture；The proliferative induction
Culture medium S2 is using MS as minimal medium, further includes: the sucrose of the IBA of the 6-BA of 0.15mg/L, 0.1mg/L, 15g/L and
The agar of 5.8g/L.
Preferably, when can only obtain green bulk callus using the proliferative induction culture medium S2, by the hair
White poplar explant is inoculated in culture on proliferative induction culture medium S3 and obtains Bud Differentiation, then the Bud Differentiation is inoculated in proliferative induction
It is cultivated on culture medium S1, obtains proliferation bud；The proliferative induction culture medium S3 is using MS as minimal medium, further includes: the sugarcane of 30g/L
The agar of sugar and 5.8g/L, pH value are 5.8~6.2.
Preferably, after carrying out root induction culture with root induction culture medium R1, low rooting rate but plant height, leaf are obtained
It further include carrying out culture of rootage using root induction culture medium R2 when piece size grows the normal Chinese white poplar tissue-cultured seedling；Institute
Root induction culture medium R2 is stated using 1/2MS culture medium as minimal medium, further includes the IBA of 0.7mg/L, the sugarcane of 10~20g/L
The agar of sugar and 4.5~8.6g/L.
Preferably, after the root induction culture 15d, the culture of rootage is carried out.
Preferably, after carrying out root induction culture with root induction culture medium R1, obtain rooting rate is low, plant height increase it is slow
It further include being cultivated using root induction culture medium R3 when the slow Chinese white poplar tissue-cultured seedling；The root induction culture medium
R3 further includes the sucrose of 10~20g/L and the agar of 4.5~8.6g/L using 1/2MS culture medium as minimal medium.
Preferably, after the root induction culture 15d, the culture is carried out.
The present invention provides a kind of tissue culture methods of Chinese white poplar, including explant differentiation and proliferation, at proliferation bud, proliferation bud lures
Two continuous process of tissue-cultured seedling of taking root to obtain are led, Initial culture process is simplified；The explant of separate sources, differing maturity can be directed to
Body has been respectively adopted the randomly selected genotype of national Chinese white poplar germplasm resource bank and Chinese white poplar is ancient in embodiments of the present invention
It sets (age of tree about 300 years) and carries out tissue culture, healthy and strong tissue-cultured seedling can be obtained, be easy to successfully obtain a large amount of tissue cultures in a short time
Seedling, easy to operate, breeding coefficient is high.
Detailed description of the invention
Fig. 1 is 1 adventitious bud inducing figure of the embodiment of the present invention；
Fig. 2 is 2 adventitious bud inducing figure of the embodiment of the present invention；
Fig. 3 is 3 rooting induction of the embodiment of the present invention and growing way figure；
Fig. 4 is 4 rooting induction of the embodiment of the present invention and growing way figure；
Fig. 5 is 5 rooting induction of the embodiment of the present invention and growing way figure.
Specific embodiment
The present invention provides a kind of Populus Tomentosa culture methods, comprising the following steps: 1) explant is woven to Populus Tomentosa, it will
The explant is inoculated in progress proliferative induction culture on proliferative induction culture medium, obtains proliferation bud；The proliferative induction culture is adopted
It is carried out with proliferative induction culture medium S1, the proliferative induction culture medium S1 is using MS culture medium as minimal medium, further includes:
The sucrose of the IBA of the 6-BA of 0.2mg/L, 0.1mg/L, 24~35g/L and the agar of 4.5~8.6g/L；The proliferative induction training
Feeding temperature is (22 ± 5) DEG C, and intensity of illumination is 1000~5000lx, light application time 12h/d；
2) the proliferation bud is inoculated in progress root induction culture on root induction culture medium, obtains Chinese white poplar tissue-cultured seedling；
The root induction culture is carried out using root induction culture medium R1, and the root induction culture medium R1 is with 1/2MS culture medium
Minimal medium further includes the IBA of 0.3mg/L, the sucrose of 10~20g/L and the agar of 4.5~8.6g/L；The root induction
The temperature of culture is (22 ± 5) DEG C, and intensity of illumination is 1000~5000lx, light application time 12h/d.
In the Populus Tomentosa culture method of the invention, the type of the explant preferably includes annotinous branch or will
The spray grown after diameter≤3cm bough water planting.In the present invention, the explant can derive from 0~300 year raw Chinese white poplar
Tree will preferably grow with annotinous branch thereon or after diameter≤3cm bough water planting when the explant derives from ancient tree
Spray as explant.There is no particular determinations for method of the present invention to the bough water planting, utilize the routine side of this field
Method.Explant of the present invention preferably includes to sterilize before carrying out proliferative induction culture, side of the present invention to the disinfection
There is no particular determinations for method, utilize the conventional disinfection method of this field.
Proliferative induction culture medium S1 of the present invention is using MS culture medium as minimal medium, further includes: the 6- of 0.2mg/L
The sucrose of the IBA of BA, 0.1mg/L, 24~35g/L and the agar of 4.5~8.6g/L.Proliferative induction culture medium S1 of the present invention
The concentration of middle sucrose is preferably 26~33g/L, more preferably 28~32g/L, most preferably 30g/L.Induction training of the present invention
The concentration for supporting agar in base S1 is preferably 5~8g/L, more preferably 5.5~6.5g/L, most preferably 5.8g/L.It is of the present invention
The pH value of proliferative induction culture medium S1 is preferably 5.8~6.2.The temperature of proliferative induction culture of the present invention be preferably (22 ±
4) DEG C, more preferably (23 ± 2) DEG C.The intensity of illumination of proliferative induction culture of the present invention is preferably 1000~5000lx, more
Preferably 1500~4000lx, most preferably 2000~3000lx.The light application time of proliferative induction culture of the present invention is preferred
For 12h/d.The time of proliferative induction culture of the present invention is preferably 25d.
In the present invention, when the explant is the spray obtained with ancient tree bough water planting, with proliferative induction culture
Before base S1 carries out proliferative induction culture, it is also preferable to include carry out Bud Differentiation culture using proliferative induction culture medium S2；It is described
Proliferative induction culture medium S2 is using MS as minimal medium, further includes: the sugarcane of the IBA of the 6-BA of 0.15mg/L, 0.1mg/L, 15g/L
The agar of sugar and 5.8g/L.The present invention cultivates 15d on the proliferative induction culture medium S2 can be obtained Bud Differentiation, the Bud Differentiation
25d is cultivated on proliferative induction culture medium S1 can obtain proliferation bud.The temperature of Bud Differentiation culture of the present invention be preferably (22 ±
4) DEG C, more preferably (23 ± 2) DEG C.The intensity of illumination of Bud Differentiation culture of the present invention is preferably 1000~5000lx, more excellent
It is selected as 1500~4500lx, most preferably 2000~3000lx.The light application time of Bud Differentiation culture of the present invention is preferably
12h/d.The time of Bud Differentiation culture of the present invention is preferably 30d.
In the present invention, when can only obtain green bulk callus using the proliferative induction culture medium S2, preferably
The genotype Chinese white poplar explant is inoculated in culture on proliferative induction culture medium S3 and obtains Bud Differentiation, then the Bud Differentiation is connect
Kind is cultivated on proliferative induction culture medium S1, obtains proliferation bud；The proliferative induction culture medium S3 is also wrapped using MS as minimal medium
Include: the sucrose of 30g/L and the agar of 5.8g/L, pH value are 5.8~6.2.The present invention trains on the proliferative induction culture medium S2
Callus can be obtained in feeding 15d, and the callus cultivates 30d on proliferative induction culture medium S3 can be obtained Bud Differentiation, by institute
State Bud Differentiation cultivated on proliferative induction culture medium S1 25d can obtain proliferation bud.The temperature of culture of the present invention is preferably (22
± 4) DEG C, more preferably (23 ± 2) DEG C.The intensity of illumination of culture of the present invention is preferably 1000~5000lx, more preferably
1500~4500lx, most preferably 2000~3000lx.The light application time of culture of the present invention is preferably 12h/d.The present invention
The time of the culture is preferably 30d.
After bud must being proliferated, the proliferation bud is inoculated in progress root induction culture on root induction culture medium by the present invention,
Obtain Chinese white poplar tissue-cultured seedling；The root induction culture with root induction culture medium R1 progress, the root induction culture medium R1 with
1/2MS culture medium is minimal medium, further includes the IBA of 0.3mg/L, the sucrose of 10~20g/L and the fine jade of 4.5~8.6g/L
Rouge；The temperature of the root induction culture is (23 ± 2) DEG C, and intensity of illumination is 1000~5000lx, light application time 12h/d.
Heretofore described root media R1 further includes the IBA of 0.3mg/L using 1/2MS culture medium as minimal medium,
The sucrose of 10~20g/L and the agar of 4.5~8.6g/L.The concentration of sucrose is preferred in rooting induction culture medium R1 of the present invention
For 12~18g/L, more preferably 14~16g/L, most preferably 15g/L.Agar in rooting induction culture medium R1 of the present invention
Concentration be preferably 5~8g/L, more preferably 5.5~6.5g/L, most preferably 5.8g/L.Root induction culture of the present invention
The pH value of base R1 is preferably 5.8~6.2.The temperature of root induction culture of the present invention is preferably (22 ± 4) DEG C, more preferably
(23±2)℃.The intensity of illumination of root induction culture of the present invention is preferably 1500~4500lx, more preferably 1800~
3500lx, most preferably 2000~3000lx.The light application time of root induction culture of the present invention is preferably 12h/d.This hair
The time of the bright root induction culture is preferably 25d.
In the present invention, after carrying out root induction culture with the root induction culture medium R1, obtain root amount it is few, growth
When the normal Chinese white poplar tissue-cultured seedling, it is also preferable to include carry out culture of rootage using root induction culture medium R2；The induction
Root media R2 further includes the IBA of 0.7mg/L, the sucrose of 10~20g/L and 4.5 using 1/2MS culture medium as minimal medium
The agar of~8.6g/L.The present invention on the root induction culture medium R1 after root induction culture 15d, can be obtained a small amount of root and
The normal Chinese white poplar tissue-cultured seedling of plant development.In rooting induction culture medium R2 of the present invention the concentration of sucrose be preferably 12~
18g/L, more preferably 14~16g/L, most preferably 15g/L.The concentration of agar in rooting induction culture medium R2 of the present invention
Preferably 5~8g/L, more preferably 5.5~6.5g/L, most preferably 5.8g/L.Root induction culture medium R2's of the present invention
PH value is preferably 5.8~6.2.The temperature of culture of rootage of the present invention is preferably (22 ± 4) DEG C, more preferably (23 ± 2) DEG C.
The intensity of illumination of culture of rootage of the present invention is preferably 1000~5000lx, more preferably 1500~4500lx, most preferably
2000~3000lx.The light application time of culture of rootage of the present invention is preferably 12h/d.The time of culture of rootage of the present invention
Preferably 25~35d.
In the present invention, after carrying out root induction culture with root induction culture medium R1, obtain that root amount is few, growth is slow
When the slow Chinese white poplar tissue-cultured seedling, it is also preferable to include cultivated using root induction culture medium R3；The root induction training
Base R3 is supported using 1/2MS culture medium as minimal medium, further includes the sucrose of 10~20g/L and the agar of 4.5~8.6g/L.This hair
It is bright preferably after the root induction culture 15d, carry out the culture.Sucrose in rooting induction culture medium R3 of the present invention
Concentration is preferably 12~18g/L, more preferably 14~16g/L, most preferably 15g/L.Rooting induction culture medium of the present invention
The concentration of agar is preferably 5~8g/L in R3, more preferably 5.5~6.5g/L, most preferably 5.8g/L.Induction of the present invention
The pH value of root media R3 is preferably 5.8~6.2.The temperature of culture of the present invention is preferably (22 ± 4) DEG C, more preferably
(23±2)℃.The intensity of illumination of culture of the present invention is preferably 1000~5000lx, more preferably 1500~4500lx, most
Preferably 2000~3000lx.The light application time of culture of the present invention is preferably 12h/d.
It is described in detail below with reference to tissue culture method of the embodiment to Chinese white poplar provided by the invention, but cannot be
They are interpreted as limiting the scope of the present invention.
Embodiment 1
Using S1 culture medium, to ' firm poplar No. 1 ' (breeding number state S-SV-PY-001-2013), ' firm poplar No. 3 ', (breeding is compiled
Number state S-SV-PY-003-2013) adventitious bud inducing is carried out, adventitious buds differentiation cultivation effect is good, as shown in Figure 1, wherein 1-1
It is obtained adventitious bud (inductivity 95%) for firm poplar No. 1 induction, 1-2 is that firm poplar No. 3 inductions obtain adventitious bud (inductivity 98%).
Embodiment 2
Adventitious bud proliferation induction is carried out to aging Chinese white poplar material, using S2 culture medium to about 300 years material agings of the age of tree
Serious ancient tree Chinese white poplar TS-1 carries out adventitious bud inducing, induces by 25d, no Bud Differentiation, and with light green bulk callus
Tissue occurs.Thus after S2 being changed to S3 culture medium, adventitious buds differentiation is normal after 30d, and without vitrification phenomenon.Induce result
As shown in Fig. 2, wherein 2-1 is the callus (adventitious bud induction frequency 0%) induced through S2,2-2 is to obtain after S3 is induced
The adventitious bud (adventitious bud induction frequency 90%) arrived.
Embodiment 3
Using R2 culture medium, to ' firm poplar No. 1 ' (breeding number state S-SV-PY-001-2013), ' firm poplar No. 3 ', (breeding is compiled
Number state S-SV-PY-003-2013) carry out rooting induction.It is cultivated by 25d, observes situation of taking root, taken root normal, and tissue-cultured seedling
Well-grown.Result is induced as shown in figure 3, wherein 3-1 is R2 to firm poplar No. 1 rooting induction (rooting rate 92.3%), 3-2 is
R2 is to firm poplar No. 3 rooting inductions (rooting rate 88.7%).
Embodiment 4
R1, R2 culture medium is respectively adopted, rooting induction is carried out to the woods source 1 (kind number 20160192) of Beijing provenance.Benefit
Rooting induction is carried out to the proliferation bud of woods source 1 with R1 culture medium, finds that the growth of tissue-cultured seedling plant height is normal, blade state is good after 25d
It is good, but rooting rate is low (rooting rate 30%).Thus culture medium R1 is changed to R2, woods source 1 is taken root normally after 25d, tissue culture
Seedling growth conditions are good.Result is induced as shown in figure 4, wherein 4-1 is rooting induction of the R1 to woods source 1,4-2 is R1 culture medium
The growth conditions in lower woods source 1,4-3 are R2 to the rooting induction in woods source 1 (rooting rate up to 88.5%).
Embodiment 5
R1, R2 culture medium is respectively adopted to scape woods No. 1 (kind number 20160186) progress rooting induction.Utilize R1 culture medium
Rooting induction is carried out to No. 1 proliferation bud of scape woods, discovery tissue-cultured seedling plant height increases abnormal (dwarfing) after 25d, but takes root and well (take root
Rate 85%).Thus culture medium R1 is changed to R3,25d background woods No. 1 takes root normally, and plant height increases obvious, tissue-cultured seedling growth
It is in good condition.Result is induced as shown in figure 5, wherein 5-1 is the rooting induction to scape woods No. 1,5-2 is scape woods 1 under R1 culture medium
Number growth conditions, 5-3 is R3 to scape woods No. 1 rooting induction (rooting rate 90%).
The present invention provides a kind of Populus Tomentosa culture method, the method does not include the Initial culture of traditional sense, but
The S1 or S2 that explant material is directly directly inoculated in addition hormone according to material aging degree after sterilization are cultivated, and are lured
It leads adventitious bud and generates proliferation bud, to carry out culture of rootage, scheme is simple, and tissue culture is high-efficient, and explant differentiation bud induction rate reaches
90%, rooting induction rate is up to 88.5%.
The above is only a preferred embodiment of the present invention, it is noted that for the ordinary skill people of the art
For member, various improvements and modifications may be made without departing from the principle of the present invention, these improvements and modifications are also answered
It is considered as protection scope of the present invention.
Claims (8)
1. a kind of Populus Tomentosa culture method, which comprises the following steps:
1) explant is woven to Populus Tomentosa, the explant is inoculated in progress proliferative induction training on proliferative induction culture medium
It supports, obtains proliferation bud；The proliferative induction culture is carried out using proliferative induction culture medium S1, and the proliferative induction culture medium S1 is with MS
Culture medium is minimal medium, further includes: the sucrose of the IBA of the 6-BA of 0.2mg/L, 0.1mg/L, 24~35g/L and 4.5~
The agar of 8.6g/L；The temperature of the proliferative induction culture is (22 ± 5) DEG C, and intensity of illumination is 1000~5000lx, when illumination
Between be 12h/d；
2) the proliferation bud is inoculated in progress root induction culture on root induction culture medium, obtains Chinese white poplar tissue-cultured seedling；It is described
Root induction culture is carried out using root induction culture medium R1, and the root induction culture medium R1 is basic with 1/2MS culture medium
Culture medium further includes the IBA of 0.3mg/L, the sucrose of 10~20g/L and the agar of 4.5~8.6g/L；The root induction culture
Temperature be (22 ± 5) DEG C, intensity of illumination be 1000~5000lx, light application time 12h/d.
2. Populus Tomentosa culture method according to claim 1, which is characterized in that the explant includes annotinous branch or will
The spray grown after diameter≤3cm bough water planting.
3. Populus Tomentosa culture method according to claim 2, which is characterized in that when the explant is with ancient tree bough water planting
It further include utilizing proliferative induction culture medium before carrying out proliferative induction culture with proliferative induction culture medium S1 when obtained spray
S2 carries out Bud Differentiation culture；The proliferative induction culture medium S2 is using MS as minimal medium, further includes: the 6- of 0.15mg/L
The sucrose of the IBA of BA, 0.1mg/L, 15g/L and the agar of 5.8g/L.
4. Populus Tomentosa culture method according to claim 3, which is characterized in that when utilization the proliferative induction culture medium S2
When can obtain green bulk callus, the Chinese white poplar explant is inoculated in cultivate on proliferative induction culture medium S3 and is divided
Change bud, then the Bud Differentiation is inoculated on proliferative induction culture medium S1 and is cultivated, obtains proliferation bud；The proliferative induction culture medium S3
Using MS as minimal medium, further includes: the sucrose of 30g/L and the agar of 5.8g/L, pH value are 5.8~6.2.
5. Populus Tomentosa culture method according to claim 1, which is characterized in that induced with root induction culture medium R1
After culture of rootage, low rooting rate but plant height are obtained, when leaf blade size grows the normal Chinese white poplar tissue-cultured seedling, further include benefit
Culture of rootage is carried out with root induction culture medium R2；The root induction culture medium R2 using 1/2MS culture medium as minimal medium,
It further include the IBA of 0.7mg/L, the sucrose of 10~20g/L and the agar of 4.5~8.6g/L.
6. Populus Tomentosa culture method according to claim 5, which is characterized in that after the root induction culture 15d, carry out
The culture of rootage.
7. Populus Tomentosa culture method according to claim 1, which is characterized in that induced with root induction culture medium R1
It further include being trained using root induction when obtaining the Chinese white poplar tissue-cultured seedling that rooting rate is low, plant height increasess slowly after culture of rootage
Base R3 is supported to be cultivated；The root induction culture medium R3 further includes 10~20g/L using 1/2MS culture medium as minimal medium
Sucrose and 4.5~8.6g/L agar.
8. Populus Tomentosa culture method according to claim 7, which is characterized in that after the root induction culture 15d, carry out
The culture.
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