CN1186116A

CN1186116A - Tissue culture for banana

Info

Publication number: CN1186116A
Application number: CN96114120A
Authority: CN
Inventors: 刘敏; 张纯花; 曾孟潜; 梁红健; 薛淮
Original assignee: Institute of Genetics of CAS
Current assignee: Institute of Genetics and Developmental Biology of CAS
Priority date: 1996-12-23
Filing date: 1996-12-23
Publication date: 1998-07-01
Anticipated expiration: 2016-12-23
Also published as: CN1062599C

Abstract

A plant tissue cultivation method for glutinous banana includes three stages of induction, merisis and growing root. A culture medium containing VC, biotin and high-concentration can sugar is used during the induction and merisis stages. Its advantages are high differentiation frequency and short growing period.

Description

Tissue culture for banana

The present invention relates to the method for plant tissue culture of glutinous banana.

The glutinous banana of Thailand is the treasure in the banana variety, unique flavor, and the fruit exquisiteness is fragrant and sweet, and is not perishable, storage tolerance; But traditional reproduction speed of tillering is very slow, and glutinous banana tillering ability can be lower than common banana, and therefore glutinous in the international market banana is seldom seen and cost an arm and a leg.

Adopt the tissue culture technique breeding plant can obtain very high proliferative speed, plant tissue culture technique is to utilize totipotency of plant cell (be each cell all potential develop into a new individual ability), take the cell mass on the plant materials, the small part of meristematic tissue or vegetative organ, by artificial preparation Different Nutrition composition and hormone regulating and controlling, make these histocytes form thousands of plantlets and preserved whole good inherited character in the parent, this method can obtain a large amount of test-tube plantlets at short notice, can in the workshop, carry out, thereby be called batch production production.Since produce seedling and be in vitro carry out and be placed between constant-temperature house under the illumination condition in, the four seasons can carry out, and carry out scale operation in few area, thereby land occupation not.If utilize tissue culture technique successfully to breed glutinous banana, the national conditions that the glutinous banana of Development and Production is had a large population and a few land for China are good developing direction.

Yet the tissue culture of glutinous banana is more a lot of greatly than common banana tissue culture difficulty, do not break up during with the culture medium culturing of common banana, and inductivity is extremely low.Therefore at home and abroad not seeing as yet up to now utilizes tissue culture technique successfully to breed the report of glutinous banana.

At the deficiencies in the prior art, the purpose of this invention is to provide a kind of can be fast, the tissue culture method of the glutinous banana of large-scale breeding.

Tissue culture for banana provided by the invention may further comprise the steps:

1, induce:

Meristematic tissue (as lateral bud, stem apex) with glutinous banana is seeded on the inducing culture.This substratum contains conventional minimum medium (the Murashige ﹠amp of MS; The Skoog substratum, hereinafter to be referred as MS), 6-benzyl purine (6-BA) 0.5-10, best 2-5 mg/litre, vitamins C (Vc) 0.5-10, best 1-2 mg/litre, naphthylacetic acid (NAA) 0.1-5, best 0.5-1 mg/litre, vitamin H 1-10, best 1-2 mg/litre and sucrose 30-60, preferably 40-50 grams per liter.After cultivating a couple of days, treat that culture expands when forming callus, promptly begins the next stage.

2, differentiation:

The 1st step gained culture is transferred on the division culture medium, this substratum contains the conventional minimum medium of MS, 6-benzyl purine (6-BA) 0.5-5, best 1-2 mg/litre, vitamins C (Vc) 1-2 mg/litre, vitamin H 1-10, best 1-2 mg/litre and sucrose 30-60, preferably 40-50 grams per liter.After cultivating a couple of days, begin to occur the banana seedlings of growing thickly, treat that seedling is long when routine is taken root the program desired height, enter following the 3rd step.

3, take root:

The neat root of the 2nd step gained seedling is taken off, be transferred to conventional root media.

After a couple of days, the seedling base portion begins to occur the short root of white, and grows complete root system very soon, a couple of days afterwards, can transplant according to a conventional method and grows up to normal banana plant.

The present invention has adopted two kinds of substratum that are different from common banana tissue culture, be inducing culture and division culture medium, there is not vitamin H in the substratum that common banana tissue culture is used, we have added the 1-5mg vitamin H in every liter of substratum, sucrose concentration is 30 grams in the substratum that the normal tissue cultivation is used, we have improved the concentration of sucrose in the substratum greatly, and added vitamins C at the brownization problem of culturing cell, the content 2-3 that has increased phytokinin than common banana inducing culture at our inducing culture aspect the use of hormone doubly, the use of vitamin H can make cell be in active state, the use of Vc can prevent cell brownization in culturing process, can be the high frequency differentiation than higher sucrose concentration more nutrition is provided, and the increase substratum is to the osmotic pressure of culture, thereby make the hormone of substratum better act on cell interior, the high-content hormone can make the meristematic tissue that is difficult for differentiation early start, thereby reach the purpose of seedling differentiation early, but the high-content hormone can not life-time service, in order to avoid mitogenetic deformity seedling, thereby only be used for inducing of initial stage, just change normal division culture medium then over to, just can obtain more glutinous banana test-tube plantlet.

Therefore use substratum of the present invention that following advantage is arranged: start early, it is fast to emerge, and synchronism is good, the differentiation frequency height, and growth cycle is short than other substratum, and Miao Qimiao is strong, well-grown, essentially no deformity seedling.

Adopt method provided by the invention just can begin in general about 15 days differentiation to occur,, can begin to occur growing thickly moving seedling about 15 days in differential period at induction period, after about seven days of the stage of taking root, as seen the short root of white after about 15 days, just can have been transplanted.And the routine propagation method of tillering generally can only be bred once in 1 year, once bred 3 to 10 strains, but the breeding of the inventive method anniversary once can be bred thousands of strains.

So method of the present invention, cultivate not only that plant speed is fast, breeding potential is high, and easy handling and scale operation.

The present invention will be further described below in conjunction with embodiment.

Embodiment

1, draw materials: get the lateral bud of the underground stem tuber of glutinous banana, water is rinsed well.

2, materials disinfection method: the lateral bud that takes off was soaked 30 seconds in 75% alcohol, put into 0.1% mercuric chloride solution and soak after 15-20 minute, on Bechtop, use aseptic water washing 3-4 time, back filter paper suck dry moisture.

3, inoculation: in ultra-clean work, with conventional aseptic technique method the banana lateral bud of wash clean on average is cut into 4 parts with scalper, each part volume makes progress shoot apical meristem at 3-5 millimeter 3, lies on the inducing culture.

This inducing culture consists of: MS+6-BA5 mg/litre+Vc 1 mg/litre+NAA 0.5 mg/litre+vitamin H 2 mg/litre+sucrose 40 grams per liters.

4, mitogenetic: when meristematic tissue is seeded on the inducing culture, just can see the bud that expands after 20 days, then it is transferred to division culture medium, its composition is

MS+6-BA2 mg/litre+Vc 1 mg/litre+vitamin H 1 mg/litre+sucrose 40 grams per liters

6, take root: (see figure 1) when the shape seedling length of growing thickly of differentiation arrives 3-4 centimetre, take off with the neat root of scissors, insert root media, this substratum consists of: MS+NAA0.3 mg/litre+sucrose 30 grams per liters+gac 0.5 grams per liter.

Culture temperature and illumination condition: 25 ± 2 ℃; Intensity of illumination is: the 1500-2000 lux.

7, transplant: long when having complete root, stem, leaf when the test tube seedling, it is transplanted in the matrix that contains 1/2 leech turfy soil and 1/2 vermiculite.

Method for transplanting: the test tube mouth is opened, opened wide bottleneck 24 hours at normal temperatures, tweezers are carefully taken out seedling from test tube, rinse well with tap water with long.

Transplanting medium is put into flowerpot, carefully plant seedling in the engagement, water permeable back covered with plastic film and open plastics film after 24 hours, took a breath 2 hours, prolong every day later on and open the plastics film time, till not covering fully, please note local atmospheric moisture and temperature, proper extension or minimizing cover time.The seedling of transplant survival is seen Fig. 2.

Claims

1, the method for plant tissue culture of glutinous banana is characterized in that being made up of following three steps:

(1) induces: the meristematic tissue of glutinous banana, be seeded on the inducing culture, this substratum contain the conventional minimum medium of MS, 6-benzyl purine 0.5-10 mg/litre, vitamins C 0.5-10 mg/litre, naphthylacetic acid 0.1-5 milliliter/liter, vitamin H 1-10 mg/litre and sucrose 30-60 grams per liter, cultivate a couple of days, treat that culture expands when forming callus, promptly begins the next stage;

(2) differentiation: the 1st step gained culture is transferred on the division culture medium, this substratum contains the conventional minimum medium of MS, 6-benzyl purine 0.5-5 mg/litre, vitamins C 1-2 mg/litre, vitamin H 1-10 mg/litre and sucrose 30-60 grams per liter, cultivate a couple of days, begin to occur the banana seedlings of growing thickly, treat that seedling is long when routine is taken root the program desired height, enter following the 3rd step;

(3) take root: the 2nd step gained seedling neat root is taken off, be transferred in the conventional root media, after a couple of days, the seedling base portion begins to occur the short root of white, and grows complete root system very soon, a couple of days afterwards, can transplant and grows up to normal banana plant.

2, method for plant tissue culture by the described glutinous banana of claim 1, it is characterized in that 6-benzyl purine content is the 2-5 mg/litre in the described inducing culture, Vitamin C content is the 1-2 mg/litre, naphthylacetic acid content is the 0.5-1 mg/litre, vitamin H content is the 1-2 mg/litre, and sucrose content is the 40-50 grams per liter.

3, the method for plant tissue culture by the described glutinous banana of claim 1 is characterized in that 6-benzyl purine content is the 1-2 mg/litre in the described mitogenetic substratum, and vitamin H content is the 1-2 mg/litre, and sucrose content is the 40-50 grams per liter.