CN106489736A - A kind of method for quickly breeding of hardwood nanmu - Google Patents
A kind of method for quickly breeding of hardwood nanmu Download PDFInfo
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- CN106489736A CN106489736A CN201610952816.3A CN201610952816A CN106489736A CN 106489736 A CN106489736 A CN 106489736A CN 201610952816 A CN201610952816 A CN 201610952816A CN 106489736 A CN106489736 A CN 106489736A
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- hardwood nanmu
- sprout
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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
Abstract
The present invention relates to a kind of method for quickly breeding of hardwood nanmu, belongs to tree propagation technical field.The present invention includes the selection and sterilizing, the sprouting of sprout, callus induction, differentiation and Multiplying culture, culture of rootage and hardening and transplanting of explant.It is an object of the invention to provide the method for quickly breeding of hardwood nanmu, is especially suitable for mass producing, good with formulation efficacy, bud ratio is high, and incubation time is short, and plant strain growth is vigorous, workable, the high advantage of using value.
Description
Technical field
The present invention relates to a kind of method for quickly breeding of arbor, it is more particularly related to a kind of hardwood nanmu is quick
Propagation method, belongs to tree propagation technical field.
Background technology
Hardwood nanmu is the evergreen megaphanerophyte of Lauraceae, belongs to Chinese Second Class Key Protected Plant, and the special product seeds of China, and has
High economic worth.Hardwood nanmu material is excellent, and texture is straight and structure is fine and closely woven, is unlikely to deform and ftractures, and has aroma, be building,
The preciousness material of furniture etc..Its timber and branches and leaves contain aromatic oil, and distillation can obtain machilus oil, be fine perfumery.Meanwhile, this kind is height
Megaphanerophyte, tree performance are graceful, and trunk is logical straight, and leaf throughout the year not at all, is that famous flower garden is viewed and admired and urban greening species.
Hardwood nanmu is extremely precious, and its speed of growth is very slow, and general hardwood nanmu is grown to foundation material and will go up a century.Not rare
Obtainable today, in order to save hardwood nanmu will not extinction, early in wildwood protected project implement before State Council's environment in 1984
In " rare endangered plants name person in servitude " that protective committee announces, hardwood nanmu is just significantly listed in wherein." the weight of in August, 1999
In point protection plant list ", hardwood nanmu is classified as protection object again.Therefore, the method for quickly breeding of hardwood nanmu is studied, not only can
Shorten the growth cycle of hardwood nanmu, accelerate its speed of becoming a useful person, can also enrich the domestic theoretical system that aspect is bred in hardwood nanmu, perfect
The protection work of hardwood nanmu.
State Intellectual Property Office discloses Publication No. CN103444531A in 2013.12.18, entitled " a kind of
The invention of rapid propagation method for tissue culture of phoebe zhennan ", the invention are related to a kind of rapid propagation method for tissue culture of phoebe zhennan, bag
Include following steps：（1）Seed selection, is put in matrix after pretreatment, immersion and sprouts, transfer culture after embryo differentiation；（2）Take children tender to sprout
Bar carries out bud inducement cultivation as explant；（3）The young tender seedling segment for taking new life carries out culture of rootage；（4）Indefinite clump bud is connect
Planting carries out Multiplying culture in proliferated culture medium；（5）The indefinite clump bud that Multiplying culture is obtained is inoculated in root media
Row culture of rootage；（6）Hardening and transplanting.The present invention organizes culture of rootage, and the training of indefinite clump bud proliferative induction using seedling stem section
Support and combine, rationally tame, circulation is drawn materials, farthest remains the complete genome group of silkwood, improve spun gold nanmu
The survival rate and quality of wood, accelerates the speed of growth of silkwood.
Above-mentioned prior art silkwood has certain difference with hardwood nanmu.In addition, first cultivating seed, then tissue cultures are carried out,
The ectogenesis time is longer；Using culture medium cause survival rate relatively low.
Content of the invention
Present invention seek to address that the problem of prior art tissue cultures, there is provided a kind of method for quickly breeding of hardwood nanmu, take
Tissue cultures, can effectively shorten the repoductive time of hardwood nanmu, improve bud ratio and survival rate, and nurturing staff.
To achieve these goals, the technical solution used in the present invention is：
A kind of method for quickly breeding of hardwood nanmu, it is characterised in that：Including following methods step：
A, the selection of explant and sterilizing
From sprout in the tender stem of hardwood nanmu as culture materials, first rinsed with running water, then rinsed with washing powder solution, then with certainly
Water is rinsed, and is then placed in 75% ethanol solution and is processed, and then again after 0.1% mercuric chloride sterilization with aseptic water washing, is used
Aseptic blotting paper suck dry moisture, after cutting the part of explant discoloration, is placed in standby in sterile petri dish；
B, the sprouting of sprout
Sprout after by sterilization treatment is seeded on WPM+NAA1.0mg/L+6-BA2.0mg/L+PVP6mg/L culture mediums, pH value
For 5.8-6.0, cultivation temperature is 23-27 DEG C, daily illumination 12 hours, intensity of illumination 1700Lx, and after inoculation, sprout sprouts, then
Form the seedling with leaf；
C, callus induction
The sprout in initial Fiber differentiation is taken, is placed in WPM+NAA2.0mg/L+6-BA4mg/L+PVP6mg/L culture mediums and is trained
Foster 14-16 days, cultivation temperature was 23-27 DEG C；
D, differentiation and Multiplying culture
The seedling that step B is obtained is cut and is transferred on WPM+NAA2.0mg/L+6-BA6.0mg/L+PVP10mg/L culture mediums,
Until forming Multiple Buds, pH value is 5.8-6.0, cultivation temperature 23-27 DEG C, daily illumination 14 hours, intensity of illumination 2000Lx,
It is within 24-26 days a growth cycle；
E, culture of rootage
The seedling formed after differentiation and increment culture is placed on Ms+NAA2.0mg/L+6-BA4.0mg/L+PVP10mg/L to take root training
On foster base, pH value is 5.8-6.0, cultivation temperature 23-27 DEG C, daily illumination 16 hours, intensity of illumination 2000Lx, 19-21 days shapes
Into whole plant；
F, hardening and transplanting
The culture medium bottle cap for having the hardwood nanmu seedling for taking root is opened, room temperature lower refining seedling took out seedling, is put in clear water after 2-4 days
The agar of root is cleaned up, in transfer to sandy soil or rich foster soil, is transplanted 43-47 days, is survived.
In step, the concentration of the washing powder solution is 5% to the present invention, described with the time rinsed with washing powder solution
For 5-6min.
In step, the time that is rinsed with running water is all 30-35min to the present invention.
In step, the time processed in described 75% ethanol solution is the 10-30 seconds to the present invention.
In stepb, after inoculation, 7-9 days sprouts sprout the present invention, form the seedling with leaf within 14-16 days.
In step C, During Callus Induction is light culture to the present invention.
In step D, growth coefficient is more than 5 to the present invention.
WPM culture medium prescriptions of the present invention are：Inorganic salts：NH4N03400mg/L、Ca(NO3)2·4H2O 556mg/
L、K2SO4990mg/L、CaCl2·2H2O 96mg/L、KH2PO4170mg/L、Na2MoO4·2H2O 0.25mg/L、MgSO4·
7H2O 370mg/L、MnSO4·H2O 22.4mg/L、ZnSO4·7H2O 8.6mg/L、CuSO4·5H2O 0.25mg/L、
FeSO4·7H2O 27.8mg/L、Na2-EDTA 37.3mg/L；Organic matter：Inositol 100mg/L, vitamin B1 1.0mg/L, cigarette
Sour 0.5mg/L, vitamin B6 0.5mg/L, glycine 2.0mg/L, sucrose 20g/L, agar 6g/L, pH are 5.2.
MS culture medium prescriptions of the present invention are：Inorganic salts:NH4N03l650mg/L、KNO31900mg/L、
CaCl2·2H2O 440mg/L、MgSO4·7H2O 370mg/L、KH2PO4170mg/L 、Na2-EDTA 37.3mg/L、
FeSO4·7H2O 27.8mg/L、H3BO36.2mg/L、MnSO4·4H2O 22.3mg/L、ZnSO4·7H2O 8.6mg/L、KI
0.83mg/L、Na2MoO4·2H2O 0.25mg/L、CuSO4·5H2O 0.025mg/L、CoCl2·6H2O 0.025mg/L；Have
Machine thing：Inositol 100mg/L, nicotinic acid 0.5mg/L, vitamin B6 0.5mg/L, vitamin B1 0.1mg/L, glycine
2.0mg/L.
The Advantageous Effects that the present invention brings：
1 it is an object of the invention to provide the method for quickly breeding of hardwood nanmu, is especially suitable for mass producing, good with formulation efficacy,
Bud ratio is high, and incubation time is short, and plant strain growth is vigorous, workable, the high advantage of using value.
2nd, the present invention directly carries out tissue culture using the sprout on excellent hardwood nanmu elite stand, undoubtedly shortens the cultivation time, and
Cost-effective.The present invention have chosen WPM culture mediums and MS culture mediums, during Initial culture, with WPM as minimal medium more
Be conducive to the differentiation of sprout and survive.And in process of rooting culture, MS culture mediums are more conducive to take root and later stage nurturing staff, from
And improve the survival rate of hardwood nanmu seedling.Rooting rate reaches more than 95%, transfer survival rate to more than 95%.
Specific embodiment
Embodiment 1
A kind of method for quickly breeding of hardwood nanmu, it is characterised in that：Including following methods step：
A, the selection of explant and sterilizing
From sprout in the tender stem of hardwood nanmu as culture materials, first rinsed with running water, then rinsed with washing powder solution, then with certainly
Water is rinsed, and is then placed in 75% ethanol solution and is processed, and then again after 0.1% mercuric chloride sterilization with aseptic water washing, is used
Aseptic blotting paper suck dry moisture, after cutting the part of explant discoloration, is placed in standby in sterile petri dish；
B, the sprouting of sprout
Sprout after by sterilization treatment is seeded on WPM+NAA1.0mg/L+6-BA2.0mg/L+PVP6mg/L culture mediums, pH value
For 5.8, cultivation temperature is 23 DEG C, daily illumination 12 hours, intensity of illumination 1700Lx, and after inoculation, sprout sprouts, and then forms band
The seedling of leaf；
C, callus induction
The sprout in initial Fiber differentiation is taken, is placed in WPM+NAA2.0mg/L+6-BA4mg/L+PVP6mg/L culture mediums and is trained
Support 14 days, cultivation temperature is 23 DEG C；
D, differentiation and Multiplying culture
The seedling that step B is obtained is cut and is transferred on WPM+NAA2.0mg/L+6-BA6.0mg/L+PVP10mg/L culture mediums,
Until forming Multiple Buds, pH value is 5.8,23 DEG C of cultivation temperature, daily illumination 14 hours, intensity of illumination 2000Lx, is within 24 days one
Individual growth cycle；
E, culture of rootage
The seedling formed after differentiation and increment culture is placed on Ms+NAA2.0mg/L+6-BA4.0mg/L+PVP10mg/L to take root training
On foster base, pH value is 5.8,23 DEG C of cultivation temperature, daily illumination 16 hours, intensity of illumination 2000Lx, forms whole plant within 19 days；
F, hardening and transplanting
The culture medium bottle cap for having the hardwood nanmu seedling for taking root is opened, room temperature lower refining seedling took out seedling, is put into handle in clear water after 2 days
The agar of root is cleaned up, and in transfer to sandy soil or rich foster soil, is transplanted 43 days, is survived.
Embodiment 2
A kind of method for quickly breeding of hardwood nanmu, it is characterised in that：Including following methods step：
A, the selection of explant and sterilizing
From sprout in the tender stem of hardwood nanmu as culture materials, first rinsed with running water, then rinsed with washing powder solution, then with certainly
Water is rinsed, and is then placed in 75% ethanol solution and is processed, and then again after 0.1% mercuric chloride sterilization with aseptic water washing, is used
Aseptic blotting paper suck dry moisture, after cutting the part of explant discoloration, is placed in standby in sterile petri dish；
B, the sprouting of sprout
Sprout after by sterilization treatment is seeded on WPM+NAA1.0mg/L+6-BA2.0mg/L+PVP6mg/L culture mediums, pH value
For 6.0, cultivation temperature is 27 DEG C, daily illumination 12 hours, intensity of illumination 1700Lx, and after inoculation, sprout sprouts, and then forms band
The seedling of leaf；
C, callus induction
The sprout in initial Fiber differentiation is taken, is placed in WPM+NAA2.0mg/L+6-BA4mg/L+PVP6mg/L culture mediums and is trained
Support 16 days, cultivation temperature is 27 DEG C；
D, differentiation and Multiplying culture
The seedling that step B is obtained is cut and is transferred on WPM+NAA2.0mg/L+6-BA6.0mg/L+PVP10mg/L culture mediums,
Until forming Multiple Buds, pH value is 6.0,27 DEG C of cultivation temperature, daily illumination 14 hours, intensity of illumination 2000Lx, is within 26 days one
Individual growth cycle；
E, culture of rootage
The seedling formed after differentiation and increment culture is placed on Ms+NAA2.0mg/L+6-BA4.0mg/L+PVP10mg/L to take root training
On foster base, pH value is 6.0,27 DEG C of cultivation temperature, daily illumination 16 hours, intensity of illumination 2000Lx, forms whole plant within 21 days；
F, hardening and transplanting
The culture medium bottle cap for having the hardwood nanmu seedling for taking root is opened, room temperature lower refining seedling took out seedling, is put into handle in clear water after 4 days
The agar of root is cleaned up, and in transfer to sandy soil or rich foster soil, is transplanted 47 days, is survived.
Embodiment 3
A kind of method for quickly breeding of hardwood nanmu, it is characterised in that：Including following methods step：
A, the selection of explant and sterilizing
From sprout in the tender stem of hardwood nanmu as culture materials, first rinsed with running water, then rinsed with washing powder solution, then with certainly
Water is rinsed, and is then placed in 75% ethanol solution and is processed, and then again after 0.1% mercuric chloride sterilization with aseptic water washing, is used
Aseptic blotting paper suck dry moisture, after cutting the part of explant discoloration, is placed in standby in sterile petri dish；
B, the sprouting of sprout
Sprout after by sterilization treatment is seeded on WPM+NAA1.0mg/L+6-BA2.0mg/L+PVP6mg/L culture mediums, pH value
For 5.9, cultivation temperature is 25 DEG C, daily illumination 12 hours, intensity of illumination 1700Lx, and after inoculation, sprout sprouts, and then forms band
The seedling of leaf；
C, callus induction
The sprout in initial Fiber differentiation is taken, is placed in WPM+NAA2.0mg/L+6-BA4mg/L+PVP6mg/L culture mediums and is trained
Support 15 days, cultivation temperature is 25 DEG C；
D, differentiation and Multiplying culture
The seedling that step B is obtained is cut and is transferred on WPM+NAA2.0mg/L+6-BA6.0mg/L+PVP10mg/L culture mediums,
Until forming Multiple Buds, pH value is 5.9,25 DEG C of cultivation temperature, daily illumination 14 hours, intensity of illumination 2000Lx, is within 25 days one
Individual growth cycle；
E, culture of rootage
The seedling formed after differentiation and increment culture is placed on Ms+NAA2.0mg/L+6-BA4.0mg/L+PVP10mg/L to take root training
On foster base, pH value is 5.9,25 DEG C of cultivation temperature, daily illumination 16 hours, intensity of illumination 2000Lx, forms whole plant within 20 days；
F, hardening and transplanting
The culture medium bottle cap for having the hardwood nanmu seedling for taking root is opened, room temperature lower refining seedling took out seedling, is put into handle in clear water after 3 days
The agar of root is cleaned up, and in transfer to sandy soil or rich foster soil, is transplanted 45 days, is survived.
Embodiment 4
A kind of method for quickly breeding of hardwood nanmu, it is characterised in that：Including following methods step：
A, the selection of explant and sterilizing
From sprout in the tender stem of hardwood nanmu as culture materials, first rinsed with running water, then rinsed with washing powder solution, then with certainly
Water is rinsed, and is then placed in 75% ethanol solution and is processed, and then again after 0.1% mercuric chloride sterilization with aseptic water washing, is used
Aseptic blotting paper suck dry moisture, after cutting the part of explant discoloration, is placed in standby in sterile petri dish；
B, the sprouting of sprout
Sprout after by sterilization treatment is seeded on WPM+NAA1.0mg/L+6-BA2.0mg/L+PVP6mg/L culture mediums, pH value
For 5.9, cultivation temperature is 24 DEG C, daily illumination 12 hours, intensity of illumination 1700Lx, and after inoculation, sprout sprouts, and then forms band
The seedling of leaf；
C, callus induction
The sprout in initial Fiber differentiation is taken, is placed in WPM+NAA2.0mg/L+6-BA4mg/L+PVP6mg/L culture mediums and is trained
Support 15 days, cultivation temperature is 26 DEG C；
D, differentiation and Multiplying culture
The seedling that step B is obtained is cut and is transferred on WPM+NAA2.0mg/L+6-BA6.0mg/L+PVP10mg/L culture mediums,
Until forming Multiple Buds, pH value is 5.9,24 DEG C of cultivation temperature, daily illumination 14 hours, intensity of illumination 2000Lx, is within 25 days one
Individual growth cycle；
E, culture of rootage
The seedling formed after differentiation and increment culture is placed on Ms+NAA2.0mg/L+6-BA4.0mg/L+PVP10mg/L to take root training
On foster base, pH value is 5.9,26 DEG C of cultivation temperature, daily illumination 16 hours, intensity of illumination 2000Lx, forms whole plant within 20 days；
F, hardening and transplanting
The culture medium bottle cap for having the hardwood nanmu seedling for taking root is opened, room temperature lower refining seedling took out seedling, is put into handle in clear water after 3 days
The agar of root is cleaned up, and in transfer to sandy soil or rich foster soil, is transplanted 46 days, is survived.
Embodiment 5
On the basis of embodiment 1-4：
Preferably, in step, the concentration of the washing powder solution is 5%, described with the time rinsed with washing powder solution is
5min.
Preferred or further, in step, the time that is rinsed with running water is all 30min.
Preferred or further, in step, the time processed in described 75% ethanol solution is 10 seconds.
Preferably, in stepb, after inoculation, 7 days sprouts sprout, and form the seedling with leaf within 14 days.
Preferably, in step C, During Callus Induction is light culture.
Preferably, in step D, growth coefficient is more than 5.
Preferably, described WPM culture medium prescriptions are：Inorganic salts：NH4N03400mg/L、Ca(NO3)2·4H2O
556mg/L、K2SO4990mg/L、CaCl2·2H2O 96mg/L、KH2PO4170mg/L、Na2MoO4·2H2O 0.25mg/L、
MgSO4·7H2O 370mg/L、MnSO4·H2O 22.4mg/L、ZnSO4·7H2O 8.6mg/L、CuSO4·5H2O 0.25mg/
L、FeSO4·7H2O 27.8mg/L、Na2-EDTA 37.3mg/L；Organic matter：Inositol 100mg/L, vitamin B1 1.0mg/L,
Nicotinic acid 0.5mg/L, vitamin B6 0.5mg/L, glycine 2.0mg/L, sucrose 20g/L, agar 6g/L, pH are 5.2.
Preferred or further, described MS culture medium prescriptions are：Inorganic salts:NH4N03l650mg/L、KNO3
1900mg/L、CaCl2·2H2O 440mg/L、MgSO4·7H2O 370mg/L、KH2PO4170mg/L 、Na2-EDTA
37.3mg/L、 FeSO4·7H2O 27.8mg/L、H3BO36.2mg/L、MnSO4·4H2O 22.3mg/L、ZnSO4·7H2O
8.6mg/L、KI 0.83mg/L、Na2MoO4·2H2O 0.25mg/L、CuSO4·5H2O 0.025mg/L、CoCl2·6H2O
0.025mg/L；Organic matter：Inositol 100mg/L, nicotinic acid 0.5mg/L, vitamin B6 0.5mg/L, vitamin B1 0.1mg/L,
Glycine 2.0mg/L.
Embodiment 6
On the basis of embodiment 1-4：
Preferably, in step, the concentration of the washing powder solution is 5%, described with the time rinsed with washing powder solution is
6min.
Preferred or further, in step, the time that is rinsed with running water is all 35min.
Preferred or further, in step, the time processed in described 75% ethanol solution is 30 seconds.
Preferably, in stepb, after inoculation, 9 days sprouts sprout, and form the seedling with leaf within 16 days.
Preferably, in step C, During Callus Induction is light culture.
Preferably, in step D, growth coefficient is more than 5.
Preferably, described WPM culture medium prescriptions are：Inorganic salts：NH4N03400mg/L、Ca(NO3)2·4H2O
556mg/L、K2SO4990mg/L、CaCl2·2H2O 96mg/L、KH2PO4170mg/L、Na2MoO4·2H2O 0.25mg/L、
MgSO4·7H2O 370mg/L、MnSO4·H2O 22.4mg/L、ZnSO4·7H2O 8.6mg/L、CuSO4·5H2O 0.25mg/
L、FeSO4·7H2O 27.8mg/L、Na2-EDTA 37.3mg/L；Organic matter：Inositol 100mg/L, vitamin B1 1.0mg/L,
Nicotinic acid 0.5mg/L, vitamin B6 0.5mg/L, glycine 2.0mg/L, sucrose 20g/L, agar 6g/L, pH are 5.2.
Preferred or further, described MS culture medium prescriptions are：Inorganic salts:NH4N03l650mg/L、KNO3
1900mg/L、CaCl2·2H2O 440mg/L、MgSO4·7H2O 370mg/L、KH2PO4170mg/L 、Na2-EDTA
37.3mg/L、 FeSO4·7H2O 27.8mg/L、H3BO36.2mg/L、MnSO4·4H2O 22.3mg/L、ZnSO4·7H2O
8.6mg/L、KI 0.83mg/L、Na2MoO4·2H2O 0.25mg/L、CuSO4·5H2O 0.025mg/L、CoCl2·6H2O
0.025mg/L；Organic matter：Inositol 100mg/L, nicotinic acid 0.5mg/L, vitamin B6 0.5mg/L, vitamin B1 0.1mg/L,
Glycine 2.0mg/L.
Embodiment 7
On the basis of embodiment 1-4：
Preferably, in step, the concentration of the washing powder solution is 5%, described with the time rinsed with washing powder solution is
5-6min.
Preferred or further, in step, the time that is rinsed with running water is all 32min.
Preferred or further, in step, the time processed in described 75% ethanol solution is 20 seconds.
Preferably, in stepb, after inoculation, 8 days sprouts sprout, and form the seedling with leaf within 15 days.
Preferably, in step C, During Callus Induction is light culture.
Preferably, in step D, growth coefficient is more than 5.
Preferably, described WPM culture medium prescriptions are：Inorganic salts：NH4N03400mg/L、Ca(NO3)2·4H2O
556mg/L、K2SO4990mg/L、CaCl2·2H2O 96mg/L、KH2PO4170mg/L、Na2MoO4·2H2O 0.25mg/L、
MgSO4·7H2O 370mg/L、MnSO4·H2O 22.4mg/L、ZnSO4·7H2O 8.6mg/L、CuSO4·5H2O 0.25mg/
L、FeSO4·7H2O 27.8mg/L、Na2-EDTA 37.3mg/L；Organic matter：Inositol 100mg/L, vitamin B1 1.0mg/L,
Nicotinic acid 0.5mg/L, vitamin B6 0.5mg/L, glycine 2.0mg/L, sucrose 20g/L, agar 6g/L, pH are 5.2.
Preferred or further, described MS culture medium prescriptions are：Inorganic salts:NH4N03l650mg/L、KNO3
1900mg/L、CaCl2·2H2O 440mg/L、MgSO4·7H2O 370mg/L、KH2PO4170mg/L 、Na2-EDTA
37.3mg/L、 FeSO4·7H2O 27.8mg/L、H3BO36.2mg/L、MnSO4·4H2O 22.3mg/L、ZnSO4·7H2O
8.6mg/L、KI 0.83mg/L、Na2MoO4·2H2O 0.25mg/L、CuSO4·5H2O 0.025mg/L、CoCl2·6H2O
0.025mg/L；Organic matter：Inositol 100mg/L, nicotinic acid 0.5mg/L, vitamin B6 0.5mg/L, vitamin B1 0.1mg/L,
Glycine 2.0mg/L.
Embodiment 8
On the basis of embodiment 1-4：
Preferably, in step, the concentration of the washing powder solution is 5%, described with the time rinsed with washing powder solution is
5-6min.
Preferred or further, in step, the time that is rinsed with running water is all 33min.
Preferred or further, in step, the time processed in described 75% ethanol solution is 15 seconds.
Preferably, in stepb, after inoculation, 7 days sprouts sprout, and form the seedling with leaf within 15 days.
Preferably, in step C, During Callus Induction is light culture.
Preferably, in step D, growth coefficient is more than 5.
Preferably, described WPM culture medium prescriptions are：Inorganic salts：NH4N03400mg/L、Ca(NO3)2·4H2O
556mg/L、K2SO4990mg/L、CaCl2·2H2O 96mg/L、KH2PO4170mg/L、Na2MoO4·2H2O 0.25mg/L、
MgSO4·7H2O 370mg/L、MnSO4·H2O 22.4mg/L、ZnSO4·7H2O 8.6mg/L、CuSO4·5H2O 0.25mg/
L、FeSO4·7H2O 27.8mg/L、Na2-EDTA 37.3mg/L；Organic matter：Inositol 100mg/L, vitamin B1 1.0mg/L,
Nicotinic acid 0.5mg/L, vitamin B6 0.5mg/L, glycine 2.0mg/L, sucrose 20g/L, agar 6g/L, pH are 5.2.
Preferred or further, described MS culture medium prescriptions are：Inorganic salts:NH4N03l650mg/L、KNO3
1900mg/L、CaCl2·2H2O 440mg/L、MgSO4·7H2O 370mg/L、KH2PO4170mg/L 、Na2-EDTA
37.3mg/L、 FeSO4·7H2O 27.8mg/L、H3BO36.2mg/L、MnSO4·4H2O 22.3mg/L、ZnSO4·7H2O
8.6mg/L、KI 0.83mg/L、Na2MoO4·2H2O 0.25mg/L、CuSO4·5H2O 0.025mg/L、CoCl2·6H2O
0.025mg/L；Organic matter：Inositol 100mg/L, nicotinic acid 0.5mg/L, vitamin B6 0.5mg/L, vitamin B1 0.1mg/L,
Glycine 2.0mg/L.
Embodiment 9
Case 1：
Choose sprout in the tender stem of 1000 hardwood nanmus, as culture materials, to be placed in after cleaning and sterilizing in 75% ethanol solution and process
The 10-30 seconds, then again after 0.1% mercuric chloride sterilization with aseptic water washing, with aseptic blotting paper suck dry moisture, cut explant
Behind the part of discoloration, by sterilization treatment after sprout be seeded in WPM+NAA1.0mg/L+6-BA2.0mg/L+PVP6mg/L culture
On base, pH value is 5.8-6.0, cultivation temperature（25±2）DEG C, daily illumination is 12 hours, and intensity of illumination 1700Lx, 8 after inoculation
Its sprout starts to sprout, and has 992 plant shapes into the seedling with leaf after 15 days.Take out band leaf seedling and be placed in WPM+NAA2.0mg/L+
Cultivate 15 days in 6-BA4mg/L+PVP6mg/L culture mediums, condition is light culture, cultivation temperature（25±2）℃.Then by seedling
Cut and transfer on WPM+NAA2.0mg/L+6-BA6.0mg/L+PVP10mg/L culture mediums, pH value is 5.8-6.0, cultivation temperature
（25±2）DEG C, daily illumination 14 hours, intensity of illumination 2000LX continue culture, and after 25 days, 986 form Multiple Buds.By Multiple Buds
It is placed on the root media of Ms+NAA2.0mg/L+6-BA4.0mg/L+PVP10mg/L, pH value is 5.8-6.0, cultivation temperature
（25±2）DEG C, daily illumination 16 hours, intensity of illumination 2000LX, after 20 days, 981 plant shapes are into whole plant.Will be equipped with 981 plants of hardwoods
The culture medium bottle cap of nanmu seedling is opened, and room temperature lower refining seedling took out seedling after 3 days, is put in clear water dry for the agar cleaning of root
Only, transfer in sandy soil or rich foster soil notes water conservation heat and moisture preserving, after transplanting 45 days, has 977 plants of hardwood nanmu seedling to survive, survive
Rate reaches 97.7%.
Case 2：
With 1 basic simlarity of case, its difference is：WPM+NAA2.0mg/L+6-BA4.0mg/L+ is selected in culture of rootage
PVP10mg/L is used as root media.Final hardwood nanmu seedling is survived 958 plants, survival rate to 95.8%.
Case 3：
With 1 basic simlarity of case, its difference is：Hardwood nanmu seedling after culture of rootage is directly transplanted to without hardening
In sandy soil or rich foster soil.Final hardwood nanmu seedling is survived 951 plants, and survival rate is 95.1%.
Claims (9)
1. a kind of method for quickly breeding of hardwood nanmu, it is characterised in that：Including following methods step：
A, the selection of explant and sterilizing
From sprout in the tender stem of hardwood nanmu as culture materials, first rinsed with running water, then rinsed with washing powder solution, then with certainly
Water is rinsed, and is then placed in 75% ethanol solution and is processed, and then again after 0.1% mercuric chloride sterilization with aseptic water washing, is used
Aseptic blotting paper suck dry moisture, after cutting the part of explant discoloration, is placed in standby in sterile petri dish；
B, the sprouting of sprout
Sprout after by sterilization treatment is seeded on WPM+NAA1.0mg/L+6-BA2.0mg/L+PVP6mg/L culture mediums, pH value
For 5.8-6.0, cultivation temperature is 23-27 DEG C, daily illumination 12 hours, intensity of illumination 1700Lx, and after inoculation, sprout sprouts, then
Form the seedling with leaf；
C, callus induction
The sprout in initial Fiber differentiation is taken, is placed in WPM+NAA2.0mg/L+6-BA4mg/L+PVP6mg/L culture mediums and is trained
Foster 14-16 days, cultivation temperature was 23-27 DEG C；
D, differentiation and Multiplying culture
The seedling that step B is obtained is cut and is transferred on WPM+NAA2.0mg/L+6-BA6.0mg/L+PVP10mg/L culture mediums,
Until forming Multiple Buds, pH value is 5.8-6.0, cultivation temperature 23-27 DEG C, daily illumination 14 hours, intensity of illumination 2000Lx,
It is within 24-26 days a growth cycle；
E, culture of rootage
The seedling formed after differentiation and increment culture is placed on Ms+NAA2.0mg/L+6-BA4.0mg/L+PVP10mg/L to take root training
On foster base, pH value is 5.8-6.0, cultivation temperature 23-27 DEG C, daily illumination 16 hours, intensity of illumination 2000Lx, 19-21 days shapes
Into whole plant；
F, hardening and transplanting
The culture medium bottle cap for having the hardwood nanmu seedling for taking root is opened, room temperature lower refining seedling took out seedling, is put in clear water after 2-4 days
The agar of root is cleaned up, in transfer to sandy soil or rich foster soil, is transplanted 43-47 days, is survived.
2. the method for quickly breeding of a kind of hardwood nanmu according to claim 1, it is characterised in that：In step, the laundry
The concentration of powder solution is 5%, and described is 5-6min with the time rinsed with washing powder solution.
3. the method for quickly breeding of a kind of hardwood nanmu according to claim 1 and 2, it is characterised in that：In step, described
The time that is rinsed with running water is all 30-35min.
4. the method for quickly breeding of a kind of hardwood nanmu according to claim 1 and 2, it is characterised in that：In step, described
The time processed in 75% ethanol solution is the 10-30 seconds.
5. the method for quickly breeding of a kind of hardwood nanmu according to claim 1, it is characterised in that：In stepb, 7- after inoculation
Sprout sprouts within 9 days, forms the seedling with leaf within 14-16 days.
6. the method for quickly breeding of a kind of hardwood nanmu according to claim 1, it is characterised in that：In step C, callus
Induction Process is light culture.
7. the method for quickly breeding of a kind of hardwood nanmu according to claim 1, it is characterised in that：In step D, growth coefficient
For more than 5.
8. the method for quickly breeding of a kind of hardwood nanmu according to claim 1, it is characterised in that：Described WPM culture mediums are matched somebody with somebody
Fang Wei：Inorganic salts：NH4N03400mg/L、Ca(NO3)2·4H2O 556mg/L、K2SO4990mg/L、CaCl2·2H2O
96mg/L、KH2PO4170mg/L、Na2MoO4·2H2O 0.25mg/L、MgSO4·7H2O 370mg/L、MnSO4·H2O
22.4mg/L、ZnSO4·7H2O 8.6mg/L、CuSO4·5H2O 0.25mg/L、FeSO4·7H2O 27.8mg/L、Na2-EDTA
37.3mg/L；Organic matter：Inositol 100mg/L, vitamin B1 1.0mg/L, nicotinic acid 0.5mg/L, vitamin B6 0.5mg/L,
Glycine 2.0mg/L, sucrose 20g/L, agar 6g/L, pH are 5.2.
9. the method for quickly breeding of a kind of hardwood nanmu according to claim 1 or 8, it is characterised in that：Described MS culture mediums
Fill a prescription and be：Inorganic salts:NH4N03l650mg/L、KNO31900mg/L、CaCl2·2H2O 440mg/L、MgSO4·7H2O
370mg/L、KH2PO4170mg/L 、Na2-EDTA 37.3mg/L、 FeSO4·7H2O 27.8mg/L、H3BO36.2mg/L、
MnSO4·4H2O 22.3mg/L、ZnSO4·7H2O 8.6mg/L、KI 0.83mg/L、Na2MoO4·2H2O 0.25mg/L、
CuSO4·5H2O 0.025mg/L、CoCl2·6H2O 0.025mg/L；Organic matter：Inositol 100mg/L, nicotinic acid 0.5mg/L,
Vitamin B6 0.5mg/L, vitamin B1 0.1mg/L, glycine 2.0mg/L.
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| CN108040715A (en) * | 2017-12-04 | 2018-05-18 | 大新县科学技术情报研究所 | A kind of mountain planting method of buerretiodendron hsienmu |
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| CN108401905B (en) * | 2018-04-10 | 2021-07-13 | 四川云辰园林科技有限公司 | Method for rapid propagation culture and annual fresh flower production of gerbera jamesonii |
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| CN108651284B (en) * | 2018-05-18 | 2021-09-14 | 安徽农业大学 | Method for inducing fast callus of machilus thunbergii |
| CN112772416A (en) * | 2021-01-28 | 2021-05-11 | 宜宾金铁红林业科技有限公司 | Tissue culture seedling raising method for phyllostachys microphylla |
| CN112772416B (en) * | 2021-01-28 | 2022-08-12 | 苏州金铁红林业科技有限公司 | Tissue culture seedling raising method for phyllostachys microphylla |
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