CN1468508A

CN1468508A - Method of preventing browning in plant tissue culture

Info

Publication number: CN1468508A
Application number: CNA021344035A
Authority: CN
Inventors: 吕复兵
Original assignee: FLORICULTURE RESEARCH INSTITUTE OF GUANGDONG ACADEMY OF AGRICULTURAL SCIENCES
Current assignee: FLORICULTURE RESEARCH INSTITUTE OF GUANGDONG ACADEMY OF AGRICULTURAL SCIENCES
Priority date: 2002-07-19
Filing date: 2002-07-19
Publication date: 2004-01-21
Anticipated expiration: 2022-07-19
Also published as: CN1207959C

Abstract

The method of preventing browning in plant tissue culture is to utilize the metabolic course of tissue culture product of non-browning plant to inhibit the browning of the easy-to-brown plant during tissue culture. The fast intermediate propagulum of the non-browning plant is cultured together with explant or culture of easy-to-brown plant, and the culture of the easy-to-brown plant is inoculated to the culture medium of 20-40 day cultured intermediate propagulum or to the culture medium re-compounded with the liquid culture medium with cultured intermediate propagulum to inhibit the browning of easy-to-brown plant during tissue culture or even to eliminate the browning.

Description

A kind of method that prevents that browning produces in the Plant Tissue Breeding

The present invention relates to a kind of method for plant tissue culture, particularly relate to a kind of method that prevents that browning produces in the Plant Tissue Breeding.

Produce browning through regular meeting in the breeding fast in Plant Tissue Breeding.This is owing to after the aldehydes matter oxidation that explant or culture cut surface are oozed out medium is become due to the crineous.Browning can produce in various degree murder by poisoning to different plant species explantation tissue or culture, and serious meeting causes culture death.At present, the measure that alleviates brownization has: (1) carries out preliminary treatment to maternal plant and explant, as explant is carried out high temperature or low temperature treatment, the pre-cultivation several days on the explant elder generation liquid medium within, and then cultivate on semisolid culturemedium: (2) select suitable culture base and condition of culture, the explant growth is vigorous under suitable culture base and condition of culture, stronger meristematic capacity is arranged, can alleviate brownization, as adjust the composition and the concentration of the inorganic elements in the medium, change cultivation temperature, illumination and pH value etc.; (3) in medium, add anti-browning agent, as oxidation resistant ascorbic acid, adsorbing active carbon of cysteine hydrochloride and citric acid and tool and polyvinylpyrrolidone (PVP); (4) shorten the subculture time, shift explant continuously, can alleviate the murder by poisoning of brownization like this.But prior art can only relatively alleviate the brownization degree in the certain plants tissue culture, can not solve brownization problem effectively.The plant of easy brownization of most tissue culture does not also have a kind of effectively anti-brownization technology, does not more have a kind of method that can eliminate browning fully.And some measure meeting side effect bad to the tissue culture deposits yields in the prior art, as explant being carried out high temperature preliminary treatment meeting explant is produced injury, add the activity that active carbon can reduce hormone in the medium at medium.Because existing anti-brownization technique effect is poor, can not solve brownization problem effectively, up to the present, the tissue culture sequencing approach that some easy brownization and brownization are poisoned very big crop to tissue culture is not set up as yet, causes the tissue culture of these crops very low with the success rate of breeding fast.And tissue culture is the unique effectively rapid proliferating ways of these plants with quick breeding.This has just had a strong impact on the speed of applying of these crop new varieties.The quick breeding of the orchid family ornamental plants has two kinds of approach, and the one, use aseptically sowing seeds, produce seedling, the 2nd, produce mitogenetic seedling by tissue culture.Because there is variation in seedling, be difficult to keep the kind of improved seeds, so famous and precious improved seeds are generally produced mitogenetic seedling (clone's seedling) by tissue culture.But ubiquity browning in the orchid tissue culture is especially serious with the Bowring cattleya tissue culture.Browning in the Bowring cattleya tissue culture is bigger to the murder by poisoning of explant, causes it not grow by normal growth, even explant death.To this, existing anti-brownization technology still can not overcome effectively, causes the Bowring cattleya new varieties not promote rapidly.

The invention provides a kind of method that prevents that browning produces in the Plant Tissue Breeding, this method can solve the brownization problem in the Plant Tissue Breeding effectively, reaches the purpose that easy brownization Plant Tissue Breeding and quick breeding success rate are improved.

The technical solution adopted for the present invention to solve the technical problems is: the present invention utilizes in the tissue culture metabolic process of the culture of brownization plant not to suppress brownization of the tissue culture of easy brownization plant.At first get the plant that does not produce browning in the tissue culture, form the middle brood body (hereinafter to be referred as middle brood body) of quick breeding through tissue culture.The red palm is a kind of Araeceae Anthurium ornamental plants, and no browning produces in whole tissue culture procedures.Its tissue culture explant can be used terminal bud, axillalry bud and young leaflet tablet.But because terminal bud and axillalry bud are difficult to thorough sterilization, so adopt young leaflet tablet to make explant usually.Young leaflet tablet is through 0.1% mercuric chloride solution sterilization about 10 minutes, after the aseptic washing 5～8 times, cuts into 1cm ²About fritter, be inoculated in callus inducing medium (PH5.8, agar 0.7% solidifies for 1/2MS+BA4mg/L+2,4-Dlmg/L+3% sucrose), induce callus about 50 days.Callus is at differential medium (MS+BA8mg/L+3% sucrose, PH5.8, agar 0.7% curing) going up cultivation formed differentiated tissues again in about 30 days, differentiated tissues is at shoot proliferation medium (MS+BA3mg/L+NAA0.2mg/L+3% sucrose again, PH5.8, agar 0.7% solidifies) go up successive transfer culture 2～3 times (30 days per generations) but the back forms the clump sprout tuber (the grow thickly massive texture of bud of a kind of tool, promptly the middle brood body of breeding) fast of fast breeding.Next gets in the tissue culture explant or the culture (hereinafter to be referred as Object of Development) that easily produces the plant of browning, and cultivate one of in the following method: cultivate Object of Development and middle brood body co-inoculation (1) in the medium of Object of Development; (2) middle brood body was cultivated 20～40 days earlier in the medium of Object of Development, brood body in the middle of removing then is again at this inoculation of medium Object of Development; (3) prepare the medium of Object of Development again with the liquid nutrient medium of brood body in the middle of cultivating, then the inoculated and cultured object.For example, MS minimal medium with not additional plant growth regulator and agar is cultivated the red palm, remove the red palm about 10 days, in this liquid nutrient medium, add the Object of Development tissue culture then and breed required growth regulator and agar curing agent fast, and replenish some nutrients again and prepare new medium, inoculated and cultured object on this medium again.Moth orchid is a kind of the orchid family ornamental plants, and the explant of its tissue culture has bennet bud, stem eye and young leaflet tablet etc., but often is to use the bennet bud.Absolutely there is serious browning in explant in the Moth orchid tissue culture.This has seriously influenced growing of bennet bud.The bud segment of Moth orchid bennet band or the bennet bud that has induced and red palm clump bud are inoculated in Moth orchid bennet bud medium (MS+BA3mg/L+3% sucrose soon together, PH5.8, agar 0.7% solidifies), the bottled medium 25ml of every cultivation inoculates the red palm clump sprout tuber about a bennet band bud segment or the bennet bud that has induced and 3～4 diameter 1cm.The browning that produces in the time of can eliminating Moth orchid bennet bud tissue culture so substantially absolutely produces serious browning and contrast (Moth orchid bennet bud is cultivated separately).

The invention has the beneficial effects as follows the not natural anti-brownization material of the middle brood body generation of the quick breeding of brownization Plant Tissue Breeding of utilization, can solve the brownization problem in the easy brownization Plant Tissue Breeding effectively.

Embodiment (1) gets the middle brood body (clump sprout tuber) of the tissue-culturing quick-propagation of the red palm of Araeceae Anthurium ornamental plants and medium (the MS+BA3mg/L+3% sucrose that Moth orchid bennet bud is incubated at Moth orchid bennet bud altogether, PH5.8, agar 0.7% solidifies) in, the bottled medium 25ml of every cultivation, inoculate the red palm clump sprout tuber about a bennet bud and 3～4 diameter 1cm, can eliminate the browning in the Moth orchid bennet bud tissue culture substantially.With red palm clump sprout tuber pre-cultivation 3～5 days on this medium, connect Moth orchid bennet bud again, better effects if earlier.

The middle brood body that embodiment (2) breeds the red palm fast is first at subculture medium (MS+BA3mg/L+NAA0.2mg/L+3% sucrose, PH5.8, agar 0.7% solidifies) go up successive transfer culture (bottled medium 25ml of every cultivation about 30 days, inoculate the red palm about 3～4 diameter 1cm from sprout tuber), remove culture afterwards, then Bowring cattleya axillalry bud explant is inoculated on this medium, can eliminates the browning of Bowring cattleya axillary bud tissue in cultivating substantially, and the axillalry bud sprouting of can taking root.And contrast (Bowring cattleya axillalry bud explant is inoculated in the fresh culture) whole brownization, and grow and suppressed, in addition dead.

Embodiment (3) Araeceae Anthurium ornamental plants is seen the leaf fancy candles lit in the bridal chamber at wedding and also do not produce browning in tissue culture procedures, its middle brood body that breeds fast is first at medium (MS+BA3mg/L+NAA0.2mg/L+3% sucrose, PH5.8, agar 0.7% solidifies) go up successive transfer culture about 30 days, remove culture afterwards, then U.S.A is preced with blue bud explant and is inoculated on this subculture medium, can eliminate the browning in the blue bud tissue culture of U.S. hat substantially, cultivate two weeks left and right sides bud and begin to sprout.And contrast (the blue bud explant of U.S. hat is inoculated in the fresh culture) whole brownization, grow and suppressed.

Claims

1. method that prevents that browning in the Plant Tissue Breeding from producing is characterized in that:

1) gets the plant that does not produce browning in the tissue culture, form the middle brood body of quick breeding through tissue culture;

2) culture of easy brownization plant and the middle brood body co-inoculation of brownization plant are not cultivated in the medium of easy brownization plant;

3) the middle brood body of brownization plant was not cultivated 20～40 days earlier in the medium of the Object of Development of easy brownization, and brood body in the middle of removing then is again at this inoculation of medium Object of Development;

4) with cultivating the medium that the liquid nutrient medium that is difficult for the middle brood body of brownization plant is mixed with the Object of Development of easy brownization again, inoculated and cultured object then.

2. a kind of method that prevents that browning in the Plant Tissue Breeding from producing according to claim 1, the plant that it is characterized in that not producing in the described tissue culture browning is the red palm of Araeceae Anthurium ornamental plants.