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In order to provide banana farmers with high-quality and inexpensive seedlings, after nearly 10 years of research and practice, the Tissue Culture Center of the Yunnan Academy of Agricultural Sciences Food Crop Research Institute has improved the technology and continuously tried to solve some of the common problems in banana planting seedlings. Exploring and researching on improving production technology, reducing production costs, and optimizing production processes has established a set of technical systems for fostering high-quality, high-efficiency, low-consumption and rapid-producing banana seedlings in order to promote the production and industrialization of high-quality banana seedlings. process.
1 Production process of banana tissue culture seedlings 1.1 Formulation of culture medium According to the test results of banana tissue culture seedlings produced by the tissue culture center of the Yunnan Academy of Agricultural Science and Food Crops for many years, three groups of essential culture medium were selected: Induced meristem Medium: MS+BA 3.5~4.5 mg/L+NAA 0.1~0.2 mg/L; Proliferation medium: MS+BA 2~3 mg/L+NAA 0.05~0.15 mg/L; Rooting medium: 1/2MS +NAA 0.1~0.2 mg/L+activated carbon 0.5%.
1.2 Culture conditions The above medium was added 2% to 3% of edible sugar, 6% to 7% of agar, the pH was adjusted to 5.5 to 5.8 before sterilization, and the medium was well-mixed, stirred well and weighed cleanly. The glass bottle was sealed and sterilized under a pressure of 1.1 kg/cm2 and 121e for 20 min, and used for cooling. The optimum growth temperature for banana tissue culture seedlings is 25-28°C, which can be adjusted as necessary, and cannot be lower than 14°C and not higher than 32°C. In the early stage of culture, the production of adventitious buds was induced by weak light; at the later stage of culture, the illumination in the tissue culture room was adjusted to 2 000~3 000 lx, and the illumination time was 12 h/d.
1.3 Induction culture The purpose of the initial culture is to obtain sterile seedlings and establish a sterile propagation system. On a sunny day, in a banana area free of traditional diseases, use a mother plant that is strong, fruiting, and yielding high, dig up the intact buds that have just exposed the ground, wash the surface clay, and remove the leaf sheaths and adventitious roots outside the buds. The washing powder is washed with water and then rinsed with tap water. Finally, the sucker is trimmed with a knife (4~5)cm×(6~7)cm (diameter×height). After being brought into the inoculation room and wiped with an alcohol swab, the pseudo-stem leaf sheath is stripped off on the hyperbacterial work bench and trimmed into a sucker with a growth point of about 3 cm×3.5 cm (diameter @ height). 75% alcohol soaked for 1 min, washed with sterile water once, then sterilized with 0.1% mercury-mercury solution for 15 min, and constantly shaking the treatment solution, and then rinsed with sterile water 4 to 5 times, with the stem tip as the center Cut into 2~4 pieces, insert the culture medium, inoculate 1 bottle, place in the culturing room, grow 35~50 days, can grow new shoots.
1.4 Subculture After the above-mentioned shoots were transferred to the medium, they were cultured under low light conditions. If conditions are suitable, substituting for 20 times 30 days can be used. Adventitious buds continue to switch, split, so that the number of buds continue to increase, to achieve the purpose of proliferation.
1.5 Strong Seedlings and Rooting When the shoots are multiplied to a predetermined number, adventitious buds less than 2.5 cm in size continue to proliferate in the medium while maintaining a certain base number. Adventitious buds higher than 2.5 cm are individually cut and transferred. Rooting medium, cultured under light conditions. After about 14 days, the roots began to grow. When the plantlets had more than two green leaves, 25 to 35 days later, when the seedlings height was about 4 cm, the seedlings could be bottled.
2 Common problems in production and solutions 2.1 Variant banana tissue seedlings are susceptible to genetic variation during subculture. The mutation rate is usually higher than that of the sucking sprouts, and the abnormal buds are various in shape (if the buds are small, dwarfed, white) , flat, brown or ball-shaped base, leaf sheath spread or hypertrophy, etc.). With the increase of culture algebra, the mutation rate gradually increases [5]. Therefore, the algebra of subculture culture cannot exceed 10 generations. In the initial culture stage, the bud season, weather, and explant materials must be selected to meet the better culture conditions; in the entire production process, the types of hormones should not be used too much, the breeding stage is simple. The best effect was to use 6BAA+low concentration of NAA, and the best effect was to use NAA or IBA alone in the rooting stage. If the hormone concentration is as low/low as possible, the principle of high 0 should not be used; the sucrose concentration is 2%~3%, and the late stage of cultivation is 2%. Appropriate; In the course of inoculation, mutant seedlings should be removed in time, and the mutation rate should be controlled within 5%. Minimize mechanical damage to the material and burns as much as possible.
2.2 Pollution Pollution is a common problem in plant tissue culture, and it is also one of the most important factors affecting the production cost of banana plantlets. It largely determines the production and economic benefits of plantlets. In theory, statistics show that if the pollution rate rises by 5%, the production cost will increase by more than 10%. The higher the pollution rate, the greater the incremental rate on this basis will be more likely to cause cross-infection, and when it is serious, it will make the entire production process difficult. Normal operation. Therefore, we must take favorable measures to minimize the pollution rate. Mainly do the following work to reduce the pollution sources: Inoculation room, buffer room, culture room and medium storage room should be fumigated with formaldehyde and potassium permanganate (5~10) B1 at least once every 2 months. New Gil sterilizing water solution for mopping floors and doors, windows, culture racks and other equipment, the dry air conditioners should also be frequently activated during the rainy season to reduce the air humidity; strict requirements of the vaccination system, the use of various vaccination tools need to be constantly sterilized, all vaccination staff Need to be trained after technical training. Disinfected clothes and hats should be worn at the time of inoculation; managers should constantly check, supervise and correct irregular operations, strengthen the concept of "sterile operation" of vaccinators, and improve the inoculation method; Before culturing, they must be carefully selected. It is determined that they are sterile seedlings before they can be further subcultured. Bacterial contaminated seedlings with light bacteria can continue to be used as rooting seedlings.
2.3 Browning bananas contain tannins and other substances in the body. In the early stage of cultivation, some phenols that leak out when exposed to oxygen in the air, the polyphenol oxidase in the tissue is activated, and the banana material is automatically oxidized. Catalyzed oxidation of phenols to the corresponding quinones. Diffusion into the medium, brownish brown, These substances inhibit the activity of other enzymes, poison cultures, affect the proliferation of adventitious buds, and severely stop the growth of cultured materials. , Loss of differentiation, and ultimately browning death. Production can be based on different actual conditions, choose different ways to reduce browning. The following measures are mainly used: Anti-phenolic antioxidants (commonly known as cysteine, ascorbic acid, dithiothreitol, glutathione, thioethanol, diethyldithiocarbamate, etc.) are formulated to be suitable. Concentration, filter sterilization, wash the wound surface of the freshly cut explant; select the best culture conditions, reduce the concentration of inorganic salts, sugars and hormones in the medium, and add the appropriate amount of activated carbon (0.5~2.0 mg/L). Reduce the temperature of the culturing chamber; adopt the method of continuous transfer and timely transfer to the fresh medium; when inoculating, strictly select the site of the explant, and the action is rapid, to avoid exposing the explant in the air for too long, Minimize mechanical damage to the explants as much as possible.
3 Discussion The production cost of banana tissue culture seedlings consists of three parts: material costs, utilities, and labor costs, of which materials and utility costs each account for about 30%, and labor costs account for about 40%. In order to reduce the production cost of non-toxic seedlings, after many years of exploration, the key features of the banana production have been improved and the two key links in the production process have been improved.
3.1 Optimization of the medium formulation The appropriate medium formulation is conducive to improving the quality of banana seedlings and reducing production costs. In the medium preparation process, tap water was used instead of distilled water, white sugar was used instead of sucrose, and agricultural type 6OBA was used instead of chemical reagent 6 OBA. The MS medium was modified to remove inositol from the original medium, reduce the trace elements in the MS to 1/2 of the original amount, reduce the amount of sugar to 3% in the rooting stage, and the amount of agar could be reduced during the proliferation stage. At 5%, it was found that there was no significant effect on the quality of tissue culture seedlings.
3.2 Increasing the level of technology and management There are many and complex factors that affect the industrial production quality and production efficiency of banana tissue culture seedlings. But technology is the foundation and management is the key.
Must continuously improve the production facilities, provide the best temperature and light control conditions, reduce the types and number of bacteria in the production workshop; strengthen scientific research and technology, continue to carry out technological innovation, improve the technical level; at the same time carry out scientific management, through economic benefits and production process Linkages, increase work enthusiasm, and optimize the distribution of labor resources. At present, there are many studies on the formulation of the culture medium, and there are few studies on its comprehensive management, and it needs to be further strengthened. Only if the technology and management are constantly improved and improved, and they are coordinated, can they optimize their production processes and reduce their The cost of production will continue to increase the economic benefits of factory-based rapid propagation of high-quality banana seedlings.
Tissue Culture of Banana Seedlings
Bananas are one of the four best fruits in South China. It is a herbaceous fruit tree with fast growing speed, short production period, high yield, and good economic benefit. It is highly valued by banana producing countries and banana farmers. Compared with traditional budding propagation, banana tissue rapid propagation has the advantages of no virus, low mutation rate, uniform growth, good fruit commerciality, and convenient transportation of seedlings. Since the promotion, the demand has been increasing, and banana tissue culture seedlings have been produced. Manufacturers are also growing. In the process of banana planting seedlings, due to the occurrence of mutations and serious pollution problems, the cost of these products increased and the commodity rate decreased, which restricted the development of large-scale banana production.