CN104782501B - May melon vine seedling cultural method - Google Patents
May melon vine seedling cultural method Download PDFInfo
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- CN104782501B CN104782501B CN201510238431.6A CN201510238431A CN104782501B CN 104782501 B CN104782501 B CN 104782501B CN 201510238431 A CN201510238431 A CN 201510238431A CN 104782501 B CN104782501 B CN 104782501B
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Abstract
The invention belongs to field of plant cell engineering technology, May, the incubation step of melon vine seedling included pretreatment, aseptic explant acquisition, callus induction, inducing clumping bud propagation, culture of rootage and transplanting successively；Using the technical scheme of present invention production melon vine seedling in May, its beneficial effect produced is：（1）Proliferation rate is high, and biological yield is big, and cultivation cycle is short；（2）Tissue-cultured seedling quality is good, and seedling is healthy and strong, and cultivation survival rate is high；The present invention by pretreatment, aseptic explant acquisitions, callus induction, inducing clumping bud using being bred and culture of rootage and the method for transplanting are cultivated melon vine seedling in May relative to other training methods with the numerous efficiency high of expansion, incubation time be short, switching is convenient；Seedling is healthy and strong, cultivation survival rate is high and cost is low, is adapted to melon vine seedling large-scale cultivation production in May.
Description
Technical field
The invention belongs to field of plant cell engineering technology, and in particular to one kind utilizes Plant Tissue Breeding mode General Office
Reason carries out the fast numerous method to obtain a large amount of seedlings of melon vine culture in May.
Background technology
May melon vine（Holboellia fargesii Reaub.）For Lardizabalaceae（Lardizabalaceae）Fructus Akebiae belongs to
（Holboellia）Plant, originates in Yunnan, Guizhou, Sichuan, Hubei, Hunan, Shaanxi, Anhui, Guangxi, Guangdong and Fujian.It is wild
It is distributed in the hillside shaw of 500-3000 meters of height above sea level and cheuch woods.May, melon vine was evergreen bejuco, flower male and female
Homophyletic, red aubergine mulberry is green white or faint yellow, the short racemes of several composition umbrella room formulas, and valency is viewed and admired with higher
Value, can be cultivated as vertical greening material；The 4-5 months at its florescence, the fruiting period 7-8 months, fruit surface purple, Long Circle, long 5-9
Centimetre, pulp is sweet, can be used as application；Its other seed oil-containing 40%, can extract oil, be used as oil crops；In addition its root
It is medicinal, epersalgia cough is controlled, lumbago due to the kidney deficiency, hernia etc. is really controlled.Melon vine in May has very high application value as can be seen here, with pole
Big potentiality to be exploited.
Because the destruction of ecological environment and the harvesting of predation formula are excavated, May, the resource of melon vine reduced increasingly, had a strong impact on
Its orderly exploitation, it is therefore desirable to the exploitation of sapling multiplication technology is carried out to it.
The culture for carrying out melon vine seedling in May using the mode of Plant Tissue Breeding numerous soon is that a kind of preferable scale is obtained
May melon vine seedling method, and this method have no report and openly apply.
The content of the invention
The technical problem to be solved in the present invention is to provide a kind of growth cycle is short, transplanting survival rate is high, cost is low, operation is simple
The cultural method of single melon vine seedling in May.
In order to solve the above technical problems, the technical solution adopted by the present invention is：Incubation step successively include pretreatment, it is sterile
Explant acquisition, induction of callus, inducing clumping bud propagation and culture of rootage and transplanting；Specifically include following steps：
（1）Pretreatment：The type of different explants is using different dipping pretreatments and refrigeration pretreatment；
（2）Aseptic explant is obtained：The sterilization treatment that carried out disinfection to explant obtains aseptic explant and trained for next step
Support；
（3）Induction of callus：By step（2）The aseptic explant of middle acquisition is seeded to the induction of liquid shallow callus
The Fiber differentiation that callus is carried out on culture medium obtains callus in good condition；
（4）Inducing clumping bud is bred：By step（3）The good callus of middle acquisition is inoculated into the double-deck Multiple Buds of solid-liquid
Adventitious buds proliferation culture is carried out on culture medium；
（5）Culture of rootage and transplanting：By step（4）Solid culture of rootage is inoculated into after the Multiple Buds immersion treatment of middle acquisition
Culture of rootage is carried out on base, then will be cultivated after obtained rooted seedling hardening and in matrix grow obtaining melon vine kind in May
Seedling.
Using above-mentioned technical proposal, confirmed by many experiments, May, the cultural method of melon vine seedling can realize increasing
Grow rate high, biological yield is big, and cultivation cycle is short；Tissue-cultured seedling quality is good, and seedling is healthy and strong, and cultivation survival rate is high.
It is as further improve of the invention, described step（1）In the explant for blade or petiole or
Edible tender branch..The explant that the present invention is used is blade or petiole or edible tender branch, can be realized by a variety of explants to five
The culture of month melon vine seedling, had both facilitated the acquisition of explant, material is saved again and avoids waste.
It is as further improve of the invention, described step（1）In, when selected explant be blade or petiole or
During edible tender branch, the method for the pretreatment is：By the explant it is in vitro after be immediately placed on 1 ~ 5g/L of addition PVP+1 ~ 3g/L
In the Vc aqueous solution carry out 3 ~ 5h of immersion treatment, after be placed in 2 ~ 8 DEG C under the conditions of moisturizing refrigerate 5 ~ 10 d.Explant is located in advance
Reason, the preprocess method used is adapted to the follow-up cultivation of each explant, and the preprocessing process is conducive to follow-up culture,
Shorten cultivation cycle.
It is as further improve of the invention, described step（2）In, when selected explant be blade or petiole or
During edible tender branch, the mode of sterilization treatment is：Explant after having pre-processed is used after 75% 20 ~ 40s of alcohol disinfecting using liter
Mercury 5 ~ 10min of sterilizing, then aseptic water washing is standby.Sterile processing be to follow-up incubation it is very crucial, directly
Inductivity, proliferation rate and the rooting rate of follow-up cultivation are influenceed, by lot of experiment validation, the sterilizing methods are to follow-up culture
It is best.
It is as further improve of the invention, described step（3）Described in callus inducing culture
It is formulated as WPM or Ms+1.0 ~ 4.0mg/L 2,4-D+0.05 ~ 0.5mg/L NAA+0.1 ~ 1.5g/L PVP+30 g/L sucrose,
pH5.6~6.0；Condition of culture is：The h/d of illumination 0,25 ± 1 DEG C of temperature；The liquid shallow thickness is 0.5 ~ 3cm.Callus
Inducing culture composition be directly connected to callus inductivity and callus quality height, in order to improve five
The inductivity of the callus of month melon vine explant, the present invention enters to the inducing culture of the callus of melon vine explant in May
Optimization is gone, has finally been determined by substantial amounts of screening test, melon vine in May is carried out using above-mentioned callus inducing medium
The induction of callus of explant, the inducing effect of callus is best.
It is as further improve of the invention, described step（4）Described in the double-deck adventitious shoots culture base of solid-liquid
The formula of fluid nutrient medium be the mg/L 6-BA+0.02 of WPM or Ms+1.0 ~ 4.0 ~ 0.5mg/L NAA+2.0mg/L KT+0.1
The g/L sucrose of ~ 1.5g/L PVP+30, pH5.6 ~ 6.0, thickness is 0.2 ~ 1cm；Under the double-deck adventitious shoots culture base of the solid-liquid
The solid medium of layer addition 6.5g/L agar, so as to form double layer culture base；Condition of culture is：Intensity of illumination 1500-
2000 lux, the h/d of illumination 10 ~ 15,25 ± 1 DEG C of temperature.Confirmed by substantial amounts of screening test, using the double-deck Multiple Buds of solid-liquid
Culture medium can significantly improve the proliferation rate of Multiple Buds and reduce its melting brown rate.
It is as further improve of the invention, described step（5）In middle culture of rootage and transplanting, it will grow thickly first
Bud is put into 1 ~ 5g/L of the addition PVP+1 ~ 3g/L Vc aqueous solution 1 ~ 2h of immersion, after be inoculated into solid root media
Row culture of rootage, the formula of the root media is the mg/L NAA+40 of mg/L IBA+0.01 of 1/2Ms+1.0 ~ 2.0 ~ 0.1
G/L sucrose, pH5.6 ~ 6.0；Condition of culture is：Intensity of illumination 1500-2000 lux, the h/d of illumination 10 ~ 15,25 ± 1 DEG C of temperature.
It is as further improve of the invention, described step（5）Mud is planted to after the rooted seedling hardening of middle acquisition：
Perlite=2 ~ 5:In 1 matrix, keep that appropriateness is sheltered from heat or light and 70% ~ 80% humidity is cultivated i.e. condition of culture and is：Intensity of illumination
1500-2000 lux, the h/d of illumination 10 ~ 15,20 ± 1 DEG C of temperature, so as to obtain melon vine seedling in May.
Using the technical scheme of present invention production melon vine seedling in May, its beneficial effect produced is：（1）Proliferation rate is high,
Biological yield is big, and cultivation cycle is short；（2）Tissue-cultured seedling quality is good, and seedling is healthy and strong, and cultivation survival rate is high；The present invention is using by pre-
Processing, aseptic explant acquisition, induction of callus, inducing clumping bud propagation and culture of rootage and the method for transplanting are to five
Month melon vine seedling, which is cultivated, has that the numerous efficiency high of expansion, incubation time be short, switching is convenient relative to other training methods；Seedling is good for
Strong, cultivation survival rate is high and cost is low, is adapted to melon vine seedling large-scale cultivation production in May.
Embodiment
Embodiment 1
One kind by explant of stem section using the integrated treatment of Plant Tissue Breeding mode progress melon vine culture in May soon it is numerous with
The method for obtaining a large amount of seedlings is comprised the following steps that：
（1）Pretreatment：Preprocess method is the water that stem section is immediately placed on to 4 g/L PVP+1.5g/L Vc of addition after in vitro
In solution carry out immersion treatment 4h, after be placed in 4 DEG C under the conditions of moisturizing refrigerate 9 d；
（2）Aseptic explant is obtained：Pre-process the stem section finished and utilize 10 min of mercuric chloride sterilizing after 75% alcohol disinfecting 30s,
Then aseptic water washing is standby；
（3）Induction of callus：Obtained sterile stem section explant is seeded to liquid shallow callus induction
Induction of callus is carried out on culture medium；Processing method is：Obtained stem section aseptic explant is cut to length for 1cm
After be seeded on liquid shallow induction of callus culture medium, culture medium prescription be Ms+3.0 mg/L 2,4-D+0.4mg/
L NAA+0.6g/L PVP+30 g/L sucrose, pH5.8；Condition of culture is：The h/d of illumination 0,25 ± 1 DEG C of temperature；Liquid shallow is cured
The thickness of injured tissue Fiber differentiation culture medium is 2cm；Callus induction rate is 90% under the conditions of this, and melting brown rate is 2%；Callus is
Yellow green；Wherein callus induction rate（%）The explant number of=generation callus/inoculation explant sum × 100%；
（4）Adventitious buds proliferation culture：Callus is inoculated on the double-deck adventitious buds proliferation culture medium of solid-liquid and grown thickly
The formula of the fluid nutrient medium on the upper strata in bud proliferative induction culture, solid-liquid bilayer adventitious buds proliferation culture medium is Ms+3.0 mg/
The g/L sucrose of L 6-BA+0.5mg/L NAA+2.0mg/L KT+0.6g/L PVP+30, pH5.8, its thickness is 0.2 ~ 1cm；Under
The film solid media of layer is the solid medium of addition 6.5g/L agar, and levels culture medium composition is unanimously；Condition of culture is：Light
According to intensity 1500-2000 lux, the h/d of illumination 12,25 ± 1 DEG C of temperature；Inducing clumping bud rate is 95% under the conditions of this, and proliferation rate is
1:6.5, melting brown rate is 2%；Wherein inducing clumping bud rate（%）=Multiple Buds the number produced/inoculation sum × 100%；
（5）Culture of rootage and transplanting：Multiple Buds are dipped into the addition 3g/L PVP+1g/L Vc aqueous solution first and located
Manage 1h, after be inoculated into solid root media and carry out culture of rootage, root media is 1/2Ms+1.5 mg/L IBA+0.1
Mg/L NAA+40 g/L sucrose, pH5.8；Condition of culture is：Intensity of illumination 1500-2000 lux, the h/d of illumination 12, temperature 25
±1℃；Rooting rate is 98%；Mud is planted to after obtained rooted seedling hardening：Perlite=3:In 1 matrix, appropriateness is kept to shelter from heat or light
（The area that shelters from heat or light is 18%）With 20 DEG C of temperature, cultivate obtaining melon vine seedling in May under 75% damp condition, survival rate is 90%.
Embodiment 2
One kind by explant of petiole using the integrated treatment of Plant Tissue Breeding mode progress melon vine culture in May soon it is numerous with
The method for obtaining a large amount of seedlings is comprised the following steps that：
（1）Pretreatment：Preprocess method is that petiole is immediately placed on into the water-soluble of 3 g/L PVP+1g/L Vc of addition after in vitro
In liquid carry out immersion treatment 4h, after be placed in 4 DEG C under the conditions of moisturizing refrigerate 7 d；
（2）Aseptic explant is obtained：Pre-process the petiole finished and utilize mercuric chloride sterilizing 5min after 75% alcohol disinfecting 30s, so
Aseptic water washing is standby afterwards；
（3）Induction of callus：Obtained sterile petiole explant is seeded to liquid shallow callus induction
Induction of callus is carried out on culture medium；Processing method is：Obtained petiole aseptic explant is cut to after 1cm length
It is seeded on liquid shallow induction of callus culture medium, culture medium prescription is Ms+2.0 mg/L 2,4-D+0.2mg/L
NAA+1.0g/L PVP+30 g/L sucrose, pH5.8；Condition of culture is：The h/d of illumination 0,25 ± 1 DEG C of temperature；Liquid shallow callus
The thickness for organizing Fiber differentiation culture medium is 1cm；Callus induction rate is 88% under the conditions of this, and melting brown rate is 6%, and callus is Huang
Green；
（4）Adventitious buds proliferation culture：Callus is inoculated on the double-deck adventitious buds proliferation culture medium of solid-liquid and grown thickly
The formula of the fluid nutrient medium on the upper strata in bud proliferative induction culture, solid-liquid bilayer adventitious buds proliferation culture medium is Ms+2.0 mg/
The g/L sucrose of L 6-BA+0.1mg/L NAA+2.0mg/L KT+0.6g/L PVP+30, pH5.8, its thickness is 0.2 ~ 1cm；Under
Layer solid medium is the solid medium of addition 6.5g/L agar, and levels culture medium composition is unanimously；Condition of culture is：Illumination
Intensity 1500-2000 lux, the h/d of illumination 12,25 ± 1 DEG C of temperature；Inducing clumping bud rate is 90% under the conditions of this, and proliferation rate is 1:
4.7 melting brown rate is 5%；
（5）Culture of rootage and transplanting：Multiple Buds are dipped into the addition 3g/L PVP+1g/L Vc aqueous solution first and located
Manage 1.5 h, after be inoculated into solid root media and carry out culture of rootage, the formula of root media is 1/2Ms+1.0 mg/L
IBA+0.1 mg/L NAA+40 g/L sucrose, pH5.8；Condition of culture is：Intensity of illumination 1500-2000 lux, the h/ of illumination 12
D, 25 ± 1 DEG C of temperature；Rooting rate under the conditions of this is 95%；Mud is planted to after obtained rooted seedling hardening：Perlite=3:1 base
In matter, appropriateness is kept to shelter from heat or light（The area that shelters from heat or light is 18%）With 20 DEG C of temperature, cultivate obtaining melon vine in May under 75% damp condition
Seedling, survival rate is 85%.
Embodiment 3
One kind by explant of blade using the integrated treatment of Plant Tissue Breeding mode progress melon vine culture in May soon it is numerous with
The method for obtaining a large amount of seedlings is comprised the following steps that：
（1）Pretreatment：Preprocess method is that blade is immediately placed on into the water-soluble of 3 g/L PVP+1g/L Vc of addition after in vitro
In liquid carry out immersion treatment 3h, after be placed in 4 DEG C under the conditions of moisturizing refrigerate 5 d.
（2）Aseptic explant is obtained：Pre-process the blade finished and utilize mercuric chloride sterilizing 5min after 75% alcohol disinfecting 30s, so
Aseptic water washing is standby afterwards.
（3）Induction of callus：Obtained aseptic blade explant is seeded to liquid shallow callus induction
Induction of callus is carried out in culture medium；Processing method is：Obtained blade aseptic explant is cut to size
For 1cm2After be seeded on liquid shallow callus inducing medium, the thickness of liquid shallow callus inducing medium is
1cm；Culture medium prescription is Ms+2.0 mg/L 2,4-D+0.2mg/L NAA+1.0g/L PVP+30 g/L sucrose, pH5.8；Training
Foster condition is：The h/d of illumination 0,25 ± 1 DEG C of temperature；Callus induction rate is 85% under the conditions of this, and melting brown rate is 10%；Callus is
Yellow green；
（4）Adventitious buds proliferation culture：Callus is inoculated on the double-deck adventitious buds proliferation culture medium of solid-liquid and grown thickly
The formula of the fluid nutrient medium on the upper strata in bud proliferative induction culture, solid-liquid bilayer adventitious buds proliferation culture medium is Ms+2.0 mg/
The g/L sucrose of L 6-BA+0.1mg/L NAA+2.0mg/L KT+0.6g/L PVP+30, pH5.8, its thickness is 0.2 ~ 1cm；Under
Layer solid medium is the solid medium of addition 6.5g/L agar, and upper strata is fluid nutrient medium, levels culture medium composition one
Cause；Condition of culture is：Intensity of illumination 1500-2000 lux, the h/d of illumination 12,25 ± 1 DEG C of temperature.Inducing clumping bud under the conditions of this
Rate is 95%, and proliferation rate is 1:4.7, melting brown rate is 2%；
（5）Culture of rootage and transplanting：Multiple Buds are dipped into the addition 3g/L PVP+1g/L Vc aqueous solution first and located
Manage 1h, after be inoculated into solid root media and carry out rooting induction, root media is 1/2Ms+1.0 mg/L IBA+0.1
Mg/L NAA+40 g/L sucrose, pH5.8；Condition of culture is：Intensity of illumination 1500-2000 lux, the h/d of illumination 12, temperature 25
±1℃；Rooting rate is 97%；Mud is planted to after obtained rooted seedling hardening：Perlite=3:In 1 matrix, appropriateness is kept to shelter from heat or light
（The area that shelters from heat or light is 18%）With 20 DEG C of temperature, cultivate obtaining melon vine seedling in May under 75% damp condition, survival rate is 89%.
Embodiment 4
One kind by explant of stem section using the integrated treatment of Plant Tissue Breeding mode progress melon vine culture in May soon it is numerous with
Obtain the method specific steps of a large amount of seedlings as described in Example 1, unlike the double-deck inducing clumping bud proliferated culture medium of solid-liquid
In supernatant liquid culture medium formula be Ms+1.0 mg/L 6-BA+0.5mg/L NAA+2.0mg/L KT+0.1g/L PVP
+ 30 g/L sucrose, pH5.8, its thickness is 0.2cm；Lower floor's solid medium is the solid medium of addition 6.5g/L agar, on
Lower floor's culture medium composition is consistent；Condition of culture is：Intensity of illumination 1500-2000 lux, the h/d of illumination 12,25 ± 1 DEG C of temperature；This
Under the conditions of inducing clumping bud rate be 82%, proliferation rate is 1:21, melting brown rate is 2%；Adventitious shoots culture rooting rate is 100%, survival rate
For more than 95%.
Embodiment 5
One kind by explant of stem section using the integrated treatment of Plant Tissue Breeding mode progress melon vine culture in May soon it is numerous with
Obtain the method specific steps of a large amount of seedlings as described in Example 1, unlike the double-deck inducing clumping bud proliferated culture medium of solid-liquid
In supernatant liquid culture medium formula be Ms+1.0 mg/L 6-BA+0.5mg/L NAA+2.0mg/L KT+0.1g/L PVP
+ 30 g/L sucrose, pH5.8, its thickness is 1cm；Lower floor's solid medium is adds the solid medium of 6.5g/L agar, up and down
Layer culture medium composition is consistent；Condition of culture is：Intensity of illumination 1500-2000 lux, the h/d of illumination 12,25 ± 1 DEG C of temperature；This
Inducing clumping bud rate is 80% under part, and proliferation rate is 1:20, melting brown rate is 2%；Adventitious shoots culture rooting rate is 100%, and survival rate is
More than 95%.
Embodiment 6
One kind by explant of stem section using the integrated treatment of Plant Tissue Breeding mode progress melon vine culture in May soon it is numerous with
The method specific steps of a large amount of seedlings are obtained as described in Example 1, except that wherein induction of callus, Multiple Buds
Ms and 1/2Ms are replaced with WPM by basal medium in Multiplying culture and root media, and inducing clumping bud rate is under the conditions of this
92%, proliferation rate is 1:6, melting brown rate is 4%；Adventitious shoots culture rooting rate is 86%, and survival rate is more than 88%.
Finally, in addition it is also necessary to it is noted that listed above is only several specific embodiments of the invention.Obviously, this hair
It is bright to be not limited to above example, there can also be many deformations.One of ordinary skill in the art can be from present disclosure
All deformations for directly exporting or associating, are considered as protection scope of the present invention.
Claims (5)
1. a kind of cultural method of melon vine in May seedling, incubation step includes pretreatment, aseptic explant acquisition, callus group successively
Knit Fiber differentiation, inducing clumping bud propagation, culture of rootage and transplanting；Characterized in that, specifically including following steps：
（1）Pretreatment：The type of different explants is using different dipping pretreatments and refrigeration pretreatment；
（2）Aseptic explant is obtained：The sterilization treatment that carried out disinfection to explant, which obtains aseptic explant, is used for next step culture；
（3）Induction of callus：By step（2）The aseptic explant of middle acquisition is seeded to liquid shallow callus Fiber differentiation
The Fiber differentiation that callus is carried out on base obtains callus in good condition；（4）Inducing clumping bud is bred：By step（3）
The good callus of middle acquisition is inoculated on the double-deck adventitious shoots culture base of solid-liquid and carries out adventitious buds proliferation culture；
（5）Culture of rootage and transplanting：By step（4）It is inoculated into after the Multiple Buds immersion treatment of middle acquisition in solid root media
Culture of rootage is carried out, then will be cultivated after obtained rooted seedling hardening and in matrix grow obtaining melon vine seedling in May；Institute
The step of stating（1）In the explant be blade or petiole or edible tender branch；Described step（1）In, the explant is pre-
The method of processing is：The explant is immediately placed on after in vitro in 1 ~ 5g/L of the addition PVP+1 ~ 3g/L Vc aqueous solution and carried out
3 ~ 5h of immersion treatment, after be placed in 2 ~ 8 DEG C under the conditions of moisturizing refrigerate 5 ~ 10 d；Described step（3）Described in culture medium formula
For WPM or MS+1.0 ~ 4.0mg/L 2,4-D+0.05 ~ 0.5mg/L NAA+0.1 ~ 1.5g/L PVP+30 g/L sucrose, pH5.6 ~
6.0；Condition of culture is：The h/d of illumination 0,25 ± 1 DEG C of temperature；The liquid shallow thickness is 0.5 ~ 3cm；Described step（4）
Described in the formula of fluid nutrient medium in the double-deck adventitious shoots culture base of solid-liquid be the mg/L 6-BA+ of WPM or MS+1.0 ~ 4.0
The g/L sucrose of 0.02 ~ 0.5mg/L NAA+2.0mg/L KT+0.1 ~ 1.5g/L PVP+30, pH5.6 ~ 6.0, thickness be 0.2 ~
1cm；The double-deck adventitious shoots culture hypobasal of the solid-liquid is the solid medium of addition 6.5g/L agar, levels culture medium composition
Unanimously, so as to form double layer culture base；Described step（5）In middle culture of rootage and transplanting, Multiple Buds are put into first
Soak 1 ~ 2h into the addition 1 ~ 5g/L PVP+1 ~ 3g/L Vc aqueous solution, after be inoculated into solid root media and taken root
Culture, the formula of the root media is the mg/L NAA+40 g/L sugarcanes of mg/L IBA+0.01 of 1/2MS+1.0 ~ 2.0 ~ 0.1
Sugar, pH5.6 ~ 6.0；Described step（5）Mud is planted to after the rooted seedling hardening of middle acquisition：Perlite=2 ~ 5:In 1 matrix,
Keep that appropriateness is sheltered from heat or light and 70% ~ 80% humidity is cultivated.
2. the cultural method of melon vine in May seedling according to claim 1, it is characterised in that described step（2）In, go out
Bacterium processing mode be：By the explant after having pre-processed use 75% 20 ~ 40s of alcohol disinfecting after using mercuric chloride sterilizing 5 ~
10min, then aseptic water washing is standby.
3. the cultural method of melon vine in May seedling according to claim 2, it is characterised in that described step（4）Middle training
Foster condition is：Intensity of illumination 1500-2000 lux, the h/d of illumination 10 ~ 15,25 ± 1 DEG C of temperature.
4. the cultural method of melon vine in May seedling according to claim 3, it is characterised in that described step（5）Middle life
In root culture and transplanting；Condition of culture is：Intensity of illumination 1500-2000 lux, the h/d of illumination 10 ~ 15,25 ± 1 DEG C of temperature.
5. the cultural method of melon vine in May seedling according to claim 4, it is characterised in that described step（5）It is middle to move
Condition of culture after cultivation is：Intensity of illumination 1500-2000 lux, the h/d of illumination 10 ~ 15,20 ± 1 DEG C of temperature, so as to obtain May
Melon vine seedling.
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