CN102657082A

CN102657082A - In-vitro culture and planting regeneration and propagation method of Xianglei honeysuckle leaves and culture medium

Info

Publication number: CN102657082A
Application number: CN2012101311827A
Authority: CN
Inventors: 于晓英; 韩蓉; 符红艳; 廖祯妮; 陈己任
Original assignee: Hunan Agricultural University
Current assignee: Hunan Agricultural University
Priority date: 2012-04-28
Filing date: 2012-04-28
Publication date: 2012-09-12
Anticipated expiration: 2032-04-28
Also published as: CN102657082B

Abstract

The invention discloses an in-vitro culture and planting regeneration and propagation method of Xianglei honeysuckle leaves and a culture medium. When being used for culturing Xianglei honeysuckle, the method and the culture medium provided by the invention can effectively solve the problems that in the existing in-vitro culture and propagation technology of the Xianglei honeysuckle, the propagation factor is low, the rooting of the obtained strains is little, the required time is long, the cost is high, the quality is low, the transplanting survival rate is low, and the market requirement on high-quality seedlings is difficult to meet.

Description

Hunan flower bud honeysuckle blade cultured in vitro and plant regeneration and expanding propagation method and medium

Technical field

The invention belongs to the plant propagation technical field, be specifically related to a kind of Hunan flower bud honeysuckle blade cultured in vitro and plant regeneration and expanding propagation method and medium.

Background technology

The former plant of Hunan flower bud honeysuckle is largeflower-like honeysuckle flower (Lonicera macranthoides Hand Mazz), is the kind of coming out from the seed selection of largeflower-like honeysuckle flower natural mutant in 1997.

In the production, the main propagation method of Hunan flower bud honeysuckle is cuttage and grafting, but cottage propagation rootage duration long (1-3 month), rooting rate not high (having only 30%～40%); With the honeysuckle is " the Hunan flower bud honeysuckle " of stock grafting, and tree crown not affine and early ageing of later stage occurs after enlarging easily, most of jaundice; What have is withered or dead, and the reproduction coefficient of two kinds of methods is all limited with reproduction speed, and low with the stripped tissue culture technology growth coefficient of existing largeflower-like honeysuckle flower; The plant that obtains takes root few, takes length, and cost is high; Quality is low, and transplanting survival rate is low, is difficult to satisfy on the market demand to its high quality seedling.

In addition, the explant that the stripped tissue culture technology of existing largeflower-like honeysuckle flower adopts mainly is stem apex, stem segment with axillary bud, and the propagation mode is the organotypic propagation mode that directly produces the bud of growing thickly from explant; The technology of inducer blade callus and differentiation again thereof relatively lags behind, and has influenced the application of modern biological skill in the seed selection of its genetic improvement and new varieties.

Summary of the invention

It is low that the present invention is intended to overcome the growth coefficient that exists in existing Hunan flower bud honeysuckle cultured in vitro and the multiplication technique; The plant that obtains takes root few, takes length, and cost is high; Quality is low; Transplanting survival rate is low, is difficult to satisfy on the market problem to the demand of its high quality seedling, and a kind of Hunan flower bud honeysuckle blade cultured in vitro and plant regeneration and expanding propagation method and medium are provided.

In order to achieve the above object, technical scheme provided by the invention is: said Hunan flower bud honeysuckle blade cultured in vitro and plant regeneration and expanding propagation method comprise the steps:

(1) inducing of blade callus: select spire or adult leaf on the flower bud honeysuckle healthy plant of Hunan; After 75% alcohol-pickled 30 seconds; With 1～2% NaClO sterilization, 6～10min, use aseptic water washing again 2～3 times then, on superclean bench, blade is cut into about 1cm ²Fritter be inoculated in the callus inducing medium, cultivated 30～40 days, the good callus of growth conditions, wherein, the inducing culture based component is: B ₅+ 6-BA 1.0mg/L+NAA0.1mg/L+3% sucrose;

(2) differentiation of calli: the callus of inducing is transferred on the differential medium, cultivated 1～2 month, obtain the more bud of growing thickly, wherein, the differentiation culture based component is: B ₅+ 6-BA 1.5mg/L+NAA0.1mg/L+3% sucrose;

(3) strong seedling culture: the intensive bud of growing thickly is separated, transfer on the 1/2MS strong seedling culture base of no hormone and cultivated for 1～2 week;

(4) culture of rootage: the unrooted that will turn out through step (3) is transferred to and carry out culture of rootage on the root media strong sprout; Can the bottle outlet acclimatization and transplants when treating the long 3-5cm of root in about 20-25 days; Wherein, the culture of rootage based component is: 1/4MS+NAA0.2mg/L+IBA0.4mg/L+1.5% sucrose.

Beneficial effect of the present invention is embodied in:

Utilize method of the present invention that Hunan flower bud honeysuckle is carried out cultured in vitro and regeneration and expand numerously, growth coefficient is high, and the differentiation of calli rate reaches 70%～80%; The 11-13 bar of taking root when the unrooted seedling rooting is cultivated 20-25 days takes weak point, seedling robust growth, and used hormone, sugar, and the concentration of MS mother liquor is also lower, provides cost savings greatly.

Embodiment

Below in conjunction with embodiment the present invention is further described.

Embodiment 1

Inducing of blade callus:

Select spire or adult leaf on the flower bud honeysuckle healthy plant of Hunan, with 75% alcohol-pickled 30 seconds after, with 2% the NaClO 6min that sterilizes, use aseptic water washing again 3 times then, on superclean bench, blade is cut into 1cm ²Fritter to be inoculated into medium component be B ₅Cultivated 30～40 days in the callus inducing medium of+6-BA 1.0mg/L+NAA 0.1mg/L+3% sucrose;

Differentiation of calli:

It is B that the callus of inducing is transferred to medium component ₅Cultivated 1～2 month the bud of must growing thickly on the differential medium of+6-BA 1.5mg/L+NAA 0.1mg/L+3% sucrose;

Strong seedling culture:

The intensive bud of growing thickly is separated, transfer on the 1/2MS strong seedling culture base of no hormone and cultivated for 1～2 week;

Culture of rootage:

Unrooted transferred to strong sprout carried out on the root media that medium component is a 1/4MS+NAA0.2mg/L+IBA0.4mg/L+1.5% sucrose culture of rootage 20-25 days, bottle outlet acclimatization and transplants when treating the long 3～5cm of root.

The effect comparison experiment of embodiment 2 Hunan of the present invention flower bud honeysuckle blade cultured in vitro and plant regeneration and expanding propagation method and prior art

Under the prerequisite of other term harmonizations, method of the present invention and cultured in vitro method of the prior art are compared experiment, experiment parameter and result are as shown in table 1:

Table 1

Can find out by table 1, compare with the existing Hunan flower bud honeysuckle tissue culture technology that exsomatizes, utilize method of the present invention and medium to Hunan flower bud honeysuckle carry out cultured in vitro and regeneration expand numerous, the growth coefficient height, the differentiation of calli rate reaches 70%～80%; The 11-13 bar of taking root when the unrooted seedling rooting is cultivated 20-25 days takes weak point, seedling robust growth, and used hormone, sugar, and the concentration of MS mother liquor is also lower, provides cost savings greatly.

Claims

1. a Hunan flower bud honeysuckle blade cultured in vitro and plant regeneration and expanding propagation method comprise the steps:

(1) inducing of blade callus: select spire or adult leaf on the flower bud honeysuckle healthy plant of Hunan, be inoculated in the callus inducing medium after the sterilization and cultivate, wherein, the inducing culture based component is: B ₅+ 6-BA 1.0mg/L+NAA0.1mg/L+3% sucrose;

(2) differentiation of calli: the callus of inducing transferred on the differential medium cultivate, wherein, the differentiation culture based component is: B ₅+ 6-BA 1.5mg/L+NAA 0.1mg/L+3% sucrose;

(3) strong seedling culture: the bud of growing thickly that will behind step (2) differentiation culture, grow separates, and transfers on the 1/2MS strong seedling culture base of no hormone and cultivates;

(4) culture of rootage: the unrooted that will turn out through step (3) is transferred to and carry out culture of rootage on the root media strong sprout, bottle outlet acclimatization and transplants then, and wherein, the culture of rootage based component is: 1/4MS+NAA0.2mg/L+IBA0.4mg/L+1.5% sucrose.

2. an inducing culture that is used for the method for claim 1 is characterized in that, said medium component is B ₅+ 6-BA 1.0mg/L+NAA0.1mg/L+3% sucrose.

3. a differential medium that is used for the method for claim 1 is characterized in that, said medium component is B ₅+ 6-BA 1.5mg/L+NAA0.1mg/L+3% sucrose.

4. a root media that is used for the method for claim 1 is characterized in that, said medium component is a 1/4MS+NAA0.2mg/L+IBA0.4mg/L+1.5% sucrose.