1. Shoot tip seedlings

Shoot tip seedlings are regenerated seedlings induced by shoot tip meristem culture of sweet potato. Some shoot tip seedlings are virus-free, but some still contain viruses. Advanced virus-free test-tube seedlings, that is, excellent test-tube seedlings confirmed by strict virus detection and strain selection, and sweet potato seedlings obtained under the conditions of rapid propagation in vitro or aphid prevention.

2. The original seed is the seed potato produced by planting advanced virus-free test-tube seedlings on the pathogen-free soil in the aphid-proof net shed. Potato seedlings bred from original potatoes are called original seedlings.

3. The original seed is the seed potato produced by using the original seedling in the field where the isolation condition exceeds 500 meters (that is, there is no common poisonous sweet potato planted within 500 meters) and the soil is free of germs. Potato seedlings bred from original seeds are called original seedlings.

4. Improved varieties are also called first-class production varieties. That is, seed potatoes produced by using the original seedlings of ordinary disease-free fields. The seedlings produced by improved varieties are good seedlings.

5. Seeds for secondary production Seeds for secondary production are seed potatoes produced by using excellent seedlings from common disease-free seed farms.

(2) Production procedure of virus-free sweet potato

The production process of virus-free sweet potato includes eight links, including excellent variety screening, shoot tip seedling raising, virus detection, excellent shoot tip seedling strain selection, rapid propagation of advanced virus-free test-tube seedlings, original seed, original seed and improved seed seedling propagation, and each link has strict requirements. The ultimate goal is to ensure the quality of seed potatoes at all levels and give full play to the yield-increasing potential of virus-free sweet potatoes.

1. Selection of fine varieties

There are many excellent varieties of sweet potato, and the yield and quality can be improved to varying degrees after detoxification. But all sweet potato varieties have certain regional adaptability and practical productivity. When detoxicating sweet potato, we must choose high-yield and high-quality varieties or special varieties suitable for large-scale planting in this area according to the climate, soil and cultivation conditions in this area. For example, Jing 553, Xu Shu 34, Sushu No.8 and Lushu No.8 are the best edible varieties in the suburbs, and Xu Shu 18, Yushu No.7, Yushu No.8, Yushu 12, Yushu 13, Lushu No.7 and Meiying 65433 should be selected as the sweet potato "three-powder" processing zone. In addition, it is particularly important to note that sweet potatoes can only remove some or some viruses from their bodies after detoxification, and their resistance to viruses, stem nematodes, root rot and other pests and diseases has not changed much. When selecting varieties, we must consider the insect-resistant characteristics of the varieties themselves. Such as Xu Shu 18 susceptible to stem nematode disease and root rot. In areas with stem nematode disease and root rot, we should try to avoid Xu Shu 18 for detoxification and demonstration and popularization. Yu Shu No.9, Yu Shu 1 1, Yu Shu 13 and Lushu No.7 should be selected for detoxification. For Henan Province, varieties such as Yushu No.7 (starch type), Jing 553 (edible type), Ji Shu No.4 (edible type) and Sushu No.8 (edible type) should be selected for root rot-free areas, and Xu Shu 18 (starch type), Yushu No.8 (starch type) and Yushu1should be selected for root rot areas. In areas with stem nematode disease and root rot, varieties such as Yushu No.9 (starch type), Yushu 13 (starch type), Yushu 1 1 (edible and vegetable type) can be selected.

2. Shoot tip seedling raising

The virus mainly spreads through vascular transport tissues, and the meristem of stem tip does not form vascular bundles. The virus is mainly transmitted through plasmodesmata, and the transmission speed is very slow. Furthermore, the metabolic activity of meristem at the top of stem is very vigorous and the concentration of growth hormone is high, so the replication of virus is greatly inhibited. Therefore, the meristem at the top of the stem carries no or little virus. Virus-free plants of sweet potato shoot tips can be regenerated by cutting the meristem of sweet potato shoot tips under aseptic conditions and culturing in vitro on a specific medium. The method of inducing shoot tip seedlings: Select 3 cm long bud segments at the top of sweet potato seedlings, disinfect them with 70% alcohol and 3% bleaching powder respectively, and peel off the shoot tip under the endoscope of clean workbench. Peeled shoot tips with a length of 0.2-0.5 mm (generally with 1-2 Yuan Ye groups) were inoculated on MS medium supplemented with 1-2 mg of 6-BA per liter, and cultured under the illumination of 26-28 degrees Celsius. After the shoot tip swells and turns green, it is transferred to hormone-free MS medium to culture into shoot tip test-tube seedlings. When the seedlings grow to 5-6 leaves, move them to a nutrition bowl for virus detection. Generally speaking, it takes at least 60-90 days from peeling the shoot tip to inducing shoot tip seedlings with 5-7 leaves. The key technology of virus-free sweet potato is to induce sweet potato shoot tip seedlings by meristem culture. Moreover, the cultivation of sweet potato shoot tip seedlings needs a tissue culture room with complete equipment, high technical level and large investment. There is no need for general units, especially grass-roots units, to carry out this research, and virus-free test-tube seedlings or original seeds confirmed by identification can be obtained from qualified units.

3. Virus detection

Not all shoot tip seedlings obtained from shoot tip meristem culture are virus-free seedlings, only some seedlings are virus-free seedlings; Shoot tip seedlings are virus-free only after strict virus detection and confirmation that they are virus-free. In the detection of shoot tip seedlings, weak seedlings and symptom seedlings were eliminated by visual inspection, and then screened by serological method or molecular biology method. Then the samples that are negative by serological or molecular biological methods are tested to indicate plant grafting.

4. Selection of excellent strains

The bud mutation rate of sweet potato is relatively high, and the shoot tip seedling lines regenerated by shoot tip meristem culture often have great differences in morphology and yield. Therefore, the virus-free vaccine confirmed by virus detection must select excellent strains, eliminate mutant strains and keep excellent strains. The method of strain selection is to plant 5- 10 virus-free seedling strain in insect-proof net room, and compare it with common virus infected potato of the same variety, observe and evaluate its morphology, growth and yield, and select some optimal strains which meet the characteristics of varieties and have high yield for mixed propagation.

5. Rapid propagation of advanced virus-free test-tube seedlings

After virus-free seedlings were obtained by shoot apex meristem culture, rapid propagation technology played a decisive role in obtaining enough virus-free seedlings for field production. The mass propagation of virus-free sweet potato shoot tip seedlings can be accomplished by rapid propagation of single leaf node of test-tube seedlings or greenhouse propagation. Both have advantages in speed and cost.

(1) Rapid propagation of virus-free test-tube seedlings has the following advantages: ① Rapid propagation. Under suitable culture conditions (temperature 25℃, light 18 hours/day), 1 stem can grow into small plants with 5-6 leaves in 1 month, and the propagation coefficient is 5n based on 5 plants 1 time. ② Avoid reinfection of virus. The rapid propagation of virus-free test-tube seedlings is carried out under strict aseptic conditions, that is, there is no virus source and no transmission medium. ③ The survival rate of subculture is high. Except for a few pollution caused by careless operation, the seedling rate of single stem and leaf reached 100%. ④ It is not limited by season, climate and space, and can be industrialized. At present, there are two rapid propagation methods of virus-free seedlings: one is liquid oscillation culture, in which a single stem node is placed in a liquid culture medium and oscillated at a speed of 80 rpm; The other is a solid culture. The advantage of the former is rapid propagation, and 20 knots of left cloth can be obtained in 15-20 days. However, due to the need to equip with a shaker, the cost is high, so the two types mostly adopt solid culture. Medium for rapid propagation of shoot tip seedlings in vitro: 1/2 medium without any hormones is generally used.

The results showed that all-MS culture medium could provide sufficient nutrients for test-tube seedlings, thus maintaining the vigorous growth of test-tube seedlings, and it was an ideal culture medium. However, from the perspective of trial propagation, under normal circumstances, the propagation coefficient mainly depends on the number of leaves and plant height. Although the growth of test-tube seedlings in L/2 MS medium is not as strong as that in complete MS medium, the above two indexes can still meet the requirements of rapid propagation, and it is also an ideal medium. In order to reduce the cost, edible white sugar can be used instead of analytical pure sucrose, and some organic components can also be subtracted. Light conditions for rapid propagation in vitro: The significance of light time to the development of sweet potato plantlets in vitro lies not only in the need of morphological development of stems and leaves, but also has a significant impact on their growth. Under the condition of dark culture, the segments of virus-free sweet potato seedlings have leaf differentiation and stem elongation, but they are yellow, and the leaves do not elongate and grow, and the stems are thin and weak. The plant height, leaf number and fresh weight of virus-free sweet potato seedlings increased with the extension of light time. Prolonging illumination time is beneficial to the differentiation and growth of stem and leaf of test-tube seedlings, and prolonging illumination time is beneficial to improving the propagation coefficient of test-tube seedlings and obtaining robust virus-free test-tube seedlings. In addition, there was no significant difference in the average internode length of virus-free sweet potato plantlets under different light hours, because the plantlets were cut into single leaf nodes for rapid propagation, and the internodes did not need to be too long. When the internode length is fixed, with the increase of the number of leaves, the propagation coefficient increases geometrically. Prolonging illumination time, with the increase of plant height, only increased the number of leaves without significantly increasing the length of internodes, which is just suitable for rapid propagation of single leaf nodes of test-tube seedlings.

(2) Rapid propagation of advanced virus-free seedlings in field

① Rapid propagation in plastic greenhouses to prevent aphids. Virus-free test-tube seedlings with 5-7 leaves were opened in mid-March, and the seedlings were tempered by room temperature illumination for 5-7 days. In the afternoon before planting, the poisonous bait mixed with 100g 40% dimethoate EC and 15-25kg dry bait was taken out from the nursery in the shed to eliminate underground pests. Then plant it in a plastic shed covered with insect-proof net according to the spacing of 5cm×5cm, cover it with a small bow shed after watering, and control the temperature at about 25 degrees Celsius. When the seedlings grow to 15-20 cm, cut off the vines and continue planting and rapid propagation. Using this double-film seedling raising method, the propagation coefficient can reach more than 100 times. However, attention should be paid to small pouring and ventilation to ensure that the temperature can neither be lower than 1O degrees Celsius nor higher than 30 degrees Celsius. ② Rapid propagation of aphid prevention net shed. That is, in late April or early May, the virus-free test-tube seedlings with 5-7 leaves after exercise are planted in an aphid-proof greenhouse at a density of 0/10,000 plants per mu, watered frequently, and peeled when the seedlings grow to about 1 5 cm to promote branching. In the future, when the branched seedlings grow to 5 leaves, continue to cut the seedlings and plant them for rapid propagation, or directly use them to propagate the original seeds. ③ Prevention and rapid propagation of aphids in warm greenhouses in winter. 65438+1At the beginning of October and the end of September, the virus-free test-tube seedlings were transplanted into a warm winter greenhouse with a 40-mesh insect net. 165438+1cover with plastic film in early October, and cover with straw curtain in early and middle February of 65438+to keep the temperature in the shed at 10-30 degrees Celsius. Pay attention to timely prevention and control of aphids. In the middle and late April, the seedlings were collected and moved to the nursery for reproduction. The exposure time of virus-free vaccine in this method is too long, the probability of reinfection is greater, and one strain is less used. It should be emphasized that sweet potato virus is mainly transmitted by Bemisia tabaci, aphids and radish aphids in a non-sustainable way. Therefore, no matter which rapid propagation method is adopted, we should remember to do isolation measures (40 mesh aphid prevention net, spatial isolation of sweet potatoes without common poison within 500 meters, etc. ) and spray aphid killing chemicals regularly to prevent aphids from spreading the virus and re-infecting.

6. Original seed breeding

The seed potato produced by advanced virus-free test-tube seedlings in the pathogen-free soil of aphid-proof net shed is the original seed. The breeding requirements of virus-free sweet potato original seeds are very strict: three conditions must be met: first, advanced virus-free test-tube seedlings must be planted. Second; It is necessary to produce the original seed in the insect-proof net shed. Insect-proof net shed is one of the important conditions for original seed breeding, and the mesh of insect-proof net used must be more than 40 meshes, so that aphids can not pass through; It can greatly reduce the probability of aphids spreading viruses. Third, the land used must be pathogen-free soil, and it is best to choose soil where sweet potatoes have not been planted for many years. In addition, it is necessary to regularly spray pesticides to control aphids and avoid re-infection of the virus. These three conditions are necessary to propagate the original species. The seed potato produced by the original seedlings or even the original seedlings planted in insect-proof net shed can not be regarded as the original seed strictly.

The original seed breeds in the insect-proof net shed, where the light is weak and the ventilation is poor, which is easy to cause flourishing growth. Therefore, when propagating the original seed, the planting density is not easy to be too high, and it is generally appropriate to have 4000 plants per mu. In management, we should pay attention to applying less nitrogen fertilizer, more phosphorus and potassium fertilizer, and controlling temperature, water and humidity to prevent stems and leaves from growing white and promote potatoes to bear more potatoes. If the virus-free sweet potato grows vigorously, the vine-lifting method can be used or 50 kilograms of water is mixed with 0.2 kilograms of potassium dihydrogen phosphate every 666.7 square meters, and the leaves are sprayed for 2-3 times to promote the swelling of sweet potato pieces and the stable growth of aboveground parts. If overgrowth occurs and the vine depth reaches more than 40 cm, it can be controlled by topping or spraying 75-150g paclobutrazol 1-2 times on the water surface of 40-70kg.

When breeding the original seed, we should always run through the consciousness of preventing the virus from reinfecting. Plant some indicator plants in the net shed and spray 1 pesticide every 1-2 weeks. The ways to control aphids are: 1.5% dimethoate powder, which is sprayed at the dosage of 1.5-2.5 kg per mu; 49% dimethoate EC 1000-2000 times, 50 kg per mu; 50% DDVP 1 500 times solution, 50 kg per mu; 50% pirimicarb wettable powder 4000 times solution, 50 kg per mu; Spray 20 ml of 20% bactericidal ester or 2.5% deltamethrin on 50 kg of water; 50% monocrotophos EC 2000-3000 times, 50 kg per mu. When controlling aphids, it is best to use a variety of chemicals in turn to prevent aphids from developing drug resistance and failing to achieve the control effect. When the original seed is harvested, it is necessary to observe whether there are virus symptoms one by one. Once the diseased strain is found, it must be resolutely removed to ensure the quality of the original seed. If the indicator plants planted in the net shed show virus symptoms, the seed potatoes propagated in the whole shed should be degraded and used.

7. Original seed breeding

Under the condition of spatial isolation of more than 500 meters, the potato pieces produced by the original seedlings (that is, the potato seedlings bred by the original seeds) are the original seeds. The propagation of the original seed must use potato seedlings as the original seedlings: it must have the space isolation conditions without common toxic sweet potato planting within 500 meters; This land must be used for at least 3 years, without ordinary poisonous sugar and without disease. Generally speaking, the number of original seeds is relatively small and the price is relatively expensive. It is best to raise seedlings as early as possible when breeding the original seeds, so as to expand the breeding area and reduce the production cost. There are many methods for rapid propagation of original seedlings, but the most commonly used methods are heating seedling raising, seedling picking in nursery and single and double leaf node propagation.

① Heating seedling raising method, also known as multistage seedling raising method. In winter or early spring (early February), we should use fire resistance, electric hotbed, double-layer plastic film covered hotbed or heated plastic greenhouse to sow in advance, strengthen management and promote the early development and rapid growth of potato seedlings. After disinfection, put the original seed potato into the seedbed. When sowing, pay attention to that the top of the potato piece is facing upwards, the potato surface is on the same plane, and the pressure between the head and tail of the seed potato should not exceed 1/3 or the potato piece should be laid flat. Remove the seam soil, pour enough bed water and cover it with plastic film. In the first week of seedling raising, the temperature of the hotbed should be around 37 degrees Celsius, and then gradually drop to 30-32 degrees Celsius. It must be noted that if the soil temperature between sowing and germination is lower than 10℃, even if the time lasts only 1 day, it will cause freezing injury, lead to potato rot and affect seedling emergence. Pay attention to uncovering the film in time after emergence to avoid burning seedlings. When potato seedlings grow to 12- 18cm with 5-7 leaves, cut the seedlings and plant them in another fire-resistant kang or hotbed to become seedlings, then cut the seedlings in two stages of seedbeds and plant them in a cold bed or greenhouse covered with a film to continue reproduction. When the temperature rises above 10 degrees Celsius in April, cuttage will be carried out and seedlings will be planted in the nursery.

(2) Nursery method is adopted.

"Chaoyanggou" nursery: the planting row spacing is 40 cm, the plant spacing is 10 cm, and the planting density is 666.7 square meters, 16000-20000 plants. Digging ditches around the seedlings in the east-west direction, planting potato seedlings close to the north of the ditch, and the seedling tips should not exceed the ridge back. This method has high leeward and sunny temperature, which is beneficial to rooting and seedling raising. Small ridge close planting nursery: ridge spacing is 50 cm, plant spacing is 10 cm, and planting density is 13000 plants/666.7m2.. Flat border nursery: make a small border with a width of 1 m, water the soles of your feet and cover them with plastic film. Transplanting seedlings according to the row spacing of 15cm× 15cm. The planting density is about 666.7 square meters, with 30,000 plants. In order to promote the growth of potato seedlings, nitrogen fertilizer can be applied in stages and properly watered. However, watering should be stopped a few days before picking seedlings to promote the robust growth of potato seedlings and improve the survival rate of seedlings.

③ Single and double leaf node propagation method. Cut 7-8 potato seedlings into short seedlings with 1 leaf and 1 leaf or 2 leaves and 2 nodes, plant them in the nursery, and plant them in the original seed breeding ground when the seedlings grow to 5-7 leaves. If single leaf nodes are used for propagation, the upper end of each node should be kept short (no more than 0.5 cm) and the lower end should be kept long. It is best to plant the seedlings after cutting them in the morning. After planting, water the nest, water it 1 time the next morning, and cover it with a layer of soil. During the breeding period, the field management should be strengthened, the management should be meticulous, and the water should be fertilized in time to promote the early emergence of buds. Except that the common poisonous sweet potato is not planted within 500m to prevent the virus transmitted by aphids from reinfecting, the other production conditions and environment for original seed reproduction are basically the same as that for common sweet potato production. Therefore, the cultivation management of the original seed breeding farm is basically the same as that of ordinary sweet potatoes. However, it is not easy to plant the original seed too early, and it is best to plant it after the end of June. In addition, the stem and leaf of virus-free sweet potato grow vigorously, so we should pay attention to controlling water and fertilizer to prevent vigorous growth. When breeding the original seed, we should pay close attention to prevent the virus from reinfecting. No common poisonous sweet potato shall be planted within 500 meters around the breeding farm, and no common poisonous sweet potato shall be planted for more than 3 years in the fields used, and there shall be no stem nematode disease, root rot and black spot disease. Plant some indicator plants in the breeding ground and spray aphid control chemicals regularly every 15 days. Observe the incidence of the virus before harvest and remove the diseased strain in time. When harvesting, strictly control the quality, and the potato pieces that do not meet the quality requirements will never enter the cellar.

8. Improve seed breeding

Potato blocks produced in ordinary fields with original seedlings (that is, buds from seedlings of original potato blocks) are called improved varieties, which are also called production seeds, that is, they are directly supplied to potato farmers to plant virus-free potato seeds. The planting, cultivation and management of improved seed breeding farm are the same as those of common sweet potato, but the fields used should be disease-free and reserved, and the management should prevent vigorous growth. If the stems and leaves of sweet potato grow too fast 40 days after potato planting and 60 days after potato planting in spring, the tendrils are too long and pale, the internodes and petioles are very long, the leaves are large and thin, and the ridges are closed prematurely, and the leaf area coefficient reaches above 3.5; Or in the middle stage of sweet potato growth, the leaves are large, the leaves are dark green, the petiole is particularly long and exceeds 2.5 times the maximum width of the leaves, the leaves are thick, the leaves are closed and airtight, and the leaf area coefficient is greater than 5, so it is considered prosperous. Specific preventive measures are:

(1) Topping 1 time at branching stage, ridge sealing stage and stem and leaf growth peak.

(2) Spraying paclobutrazol (75g to 50kg of water) 1-2 times every 666.7m2 after ridge closure. (3) When Teemo found that the growth was vigorous, Teemo immediately released Teemo 1-2 times, and each time the growth could be delayed by about 7 days. According to the research results of Xuzhou Sweet Potato Research Center (1992) in Jiangsu Province, the yield of virus-free sweet potato can be significantly increased in the first and second years, but the yield of virus-resistant varieties such as Xu Shu 18 decreased by 6. 1% in the third year, and the yield of susceptible varieties such as Liqun No.2 and Xindazi decreased by 33.6%. Therefore, after virus-free sweet potato was used continuously for 2 years, the virus reinfected seriously and the yield-increasing effect decreased. It is best to replace 1 new seed potato every two years.

(3) Seed supply procedures for virus-free sweet potato breeding.

After two years of utilization of improved varieties, the effect of increasing production is no longer obvious; New virus-free potato seeds need to be replaced 1 time every two years. Therefore, it is necessary to establish a virus-free sweet potato breeding and supply system according to the local production and economic conditions, continuously provide excellent virus-free sweet potato vines for production, and ensure the maximum yield-increasing potential of virus-free sweet potatoes.

The technology of shoot tip culture, virus detection and virus-free test-tube seedling cutting and rapid propagation in virus-free potato breeding and supply system is complicated, which requires relatively perfect tissue culture room and virus detection room, as well as specialized technicians and instruments and equipment engaged in tissue culture and virus detection. The investment is relatively large and the technology content is high, which is not suitable for prefecture-level units. It is best to concentrate funds and personnel to establish a provincial virus-free and virus detection base; The breeding of the original seed needs to invest in building a greenhouse with a 40-mesh insect net. The investment should not be too large, and it can be carried out in areas, cities or county-level units with good conditions.

The breeding of original seeds and improved seeds requires a 500-meter space isolation zone (that is, no poisonous potatoes are planted within 500 meters) and disease-free fields, with less investment, less risk and great benefit, which is suitable for county, township and village units. Generally, 65,438+0,000 advanced virus-free test-tube seedlings are introduced to the nursery in spring (early April). In summer (late June), the insect-proof net shed can breed 666.7 square meters of original seeds and harvest the original seeds 1 0,000-11,500 kg. In the early spring of the following year, the original seed potato was used for seedling raising and rapid propagation. Under the isolation condition, the original seed potato can breed 100 * 666.7 square meters and harvest the original seed potato 100-200 tons. In the third year, improved varieties can be bred 5000 * 666.7 square meters; In the fourth year, the potato seed potato with the area of 105 * 666.7 square meters can be supplied for field production. All localities can refer to this ratio and adjust it appropriately according to the technical level of local sweet potato seedling raising and production to meet the needs of local production for virus-free sweet potato seeds.

Special attention should be paid to prevent virus reinfection in the process of virus-free sweet potato reproduction. The specific measures are to cover a 40-mesh insect-proof shed, isolate the space above 500 meters, and spray aphid-killing chemicals regularly. In addition, it is best to cultivate original seeds and original seeds in the stubble where sweet potatoes have not been planted for more than 3 years, and produce seed potatoes in diseases (root rot, nematode disease, etc.). )