CN111713331A - Efficient rot disease-resistant early-stage selective breeding method for pear trees - Google Patents

Abstract

The invention relates to a pear tree high-efficiency rot-resistant early-stage selective breeding method which comprises the steps of seed selection, hybrid pollination, management of hybrid fruits, harvesting and cold storage of the hybrid fruits, seed collection, seed lamination treatment, strong seedling cultivation, seedling stage pruning, preparation before inoculation, first investigation, second investigation, evaluation and elimination of single plant disease resistance, burning of eliminated plants and single plant switching.

Description

Efficient rot disease-resistant early-stage selective breeding method for pear trees

Technical Field

The invention belongs to the technical field of breeding, and particularly relates to an efficient rot disease-resistant early-stage selective breeding method for pear trees.

Background

The pear tree rot (pear Valsa canker) is commonly called as 'rotten skin disease', is one of the most serious diseases in pear cultivation production, mainly damages a main trunk, a main branch and a side branch of a pear tree, causes bark rot, tree vigor weakness and pear fruit yield and quality reduction, has the characteristics of wide occurrence area, high morbidity and difficulty in control, and seriously attacks pear gardens, tree scabs are tired, branches and trunks are incomplete, even a large number of dead trees or garden damages are caused, and the healthy development of the pear industry in China is seriously threatened.

The pear is relatively wide in the cultivation area of China, is one of the fruit tree species which are the widest in cultivation distribution of China, and is one of the fruit trees which are deeply loved by consumers, along with the development of the whole society and economy, the requirement of people on the pear is higher and higher, the appearance is required to be attractive, the quality is required to meet the requirements of different consumer groups, the variety breeding is the root of improving the variety, more and more attention is paid to the variety breeding, the appearance is attractive, the quality is excellent, the resistance is strong, the breeding method is a main breeding target, the breeding means comprises bud change seed selection, actual seed selection, cross breeding, mutation breeding and the like, at present, 72% of new edible varieties cultivated by cross breeding in China are used as main methods and ways for breeding new pear varieties.

The process of the hybrid breeding comprises the steps of making a breeding plan according to breeding targets, collecting pollen, pollinating, bagging and isolating, managing hybrid fruits, picking the hybrid fruits in autumn, taking seeds after the hybrid fruits are refrigerated, cleaning, drying and storing the seeds, performing hybrid seed lamination treatment, sowing, maintaining in seedling stage, planting or directly taking terminal buds for grafting, starting to obtain the hybrid seedlings after 5-7 years of field management, performing comprehensive evaluation on the fruits and the like, breeding excellent plants, checking, deciding and examining to obtain a variety.

As the pear trees are highly heterozygous genetically, hybrid progeny has wide separation, and an ideal variety with multiple excellent characters needs to be obtained, a large hybrid population needs to be configured for realization, meanwhile, the childhood period of the pear trees is long, the period from sowing of hybrid seeds to the beginning of fruiting is usually 5-7 years, and the field growth of 5-7 years is realized, so that the pear trees not only have large floor area and much consumed manpower and material resources, but also have the advantages of shortening the breeding period and improving the breeding efficiency, the pre-selection research on the early characters of the fruit tree hybrid progeny is always a focus of attention of breeders, if single plants which can become excellent varieties in the future can be selected in the seedling stage, the single plants without target characters or poor characters are eliminated, and the breeding cost is greatly saved.

The resistance difference of the rot disease among different varieties is large, the possibility is provided for selecting the rot disease resistant pear varieties through breeding, the reduction of the rot disease occurrence by utilizing the resistant varieties is the most effective and safe measure for preventing and treating the rot disease in production, the resistance breeding is the key point of breeding in various countries in the world at present, and the disease resistance breeding has wide development prospect.

Disclosure of Invention

The invention aims to overcome the defects of the prior art and provide an efficient early-stage selection breeding method for pear trees with attractive appearance, excellent quality and strong resistance to rot.

The technical scheme of the invention is as follows: an efficient early-stage selective breeding method for pear trees to resist rot disease, which comprises the following steps:

step 1: selecting seeds, wherein the resistance of the filial generation to rot disease is influenced by the parent variety, the resistance of the parent variety has larger influence on the number of resistant plants of the filial generation, the number of the disease-resistant plants in the filial generation with parents not being the resistant variety is small, one of the parents or the parents is the disease-resistant variety, and more disease-resistant plants can be generated in the filial generation, so that the selected seeds are combined into one of the parents or the parents which is the disease-resistant variety, wherein the selected seeds are any two combinations of autumn pears, Chinese pears, white pears and Xinjiang pears;

step 2: performing hybrid pollination, collecting buds in a big-balloon period in advance, stripping anthers, drying in an environment at 25 ℃ after collection, sealing in an environment at 4 ℃ after pollen scattering, temporarily storing until pollination is started in a big-balloon period of female parent buds, immediately bagging and isolating after pollination, and removing the isolation after pollination for 10 days;

and step 3: management of hybrid fruits, bagging fruits three weeks after pollination, spraying insecticide and bactericide once a week before bagging, carrying out insecticide and sterilization by adopting a solution of 800 times of thiophanate methyl 70% and a solution of 4000 times of abamectin 5%, soaking bag openings by using 500 times of thiophanate methyl and 1000 times of chlorpyrifos one night before bagging, firstly propping up the bag openings when bagging, propping up the bag bottoms, expanding the bag bodies, sealing the bag openings around fruit stalks after bagging fruits, carrying out pest control in the fruit growth period, strengthening fertilizer and water management, and paying attention to fruit-strengthening fertilizer for 5 months, wherein the fertilizer mainly comprises organic fertilizer, nitrogen fertilizer and potassium fertilizer, 30 kg of urea, 30 kg of potassium sulfate and 4000 kg of biogas slurry and water per mu is applied at this time;

and 4, step 4: harvesting the hybrid fruits, starting harvesting after the fruits are fully ripe, transporting the harvested fruits back to the room to allow the harvested fruits to emit field heat, removing fruit bags, and marking;

and 5: harvesting and refrigerating, wherein in order to fully after-ripen the seeds, the hybrid fruits are harvested and radiated, then precooled for 24 hours in the environment of about 20 ℃, and then stored in a refrigeration house at 0-2 ℃;

step 6: collecting seeds, refrigerating for 60-80 days, taking the fruits out of a refrigerator, transversely cutting the fruits, taking the seeds out of ventricles by using tweezers, collecting the seeds by using seed bags, rubbing and washing the seeds at 25 ℃ in warm water, drying the seeds at a ventilated position after washing, fully drying the seeds, marking and storing at room temperature;

and 7: seed stratification treatment: laminating seeds before and after the annual New year's day, putting the seeds in warm water at 25 ℃ to fully absorb water for 12 hours, preparing a preservation box, putting fully-absorbed filter paper at the bottom of the box, then uniformly and non-overlapping the seeds on the filter paper, checking once every 10 days, and after 70-80% of the seeds are exposed to the white, starting to prepare for sowing;

and 8: cultivating strong seedlings, sowing the seeds after exposure to the white to a hole tray special for the forest with the depth of 11cm, carrying out insect killing and sterilization treatment on a matrix before sowing, starting transplanting when the seedlings grow to 4 real leaves and the roots are close to 10cm, transplanting the seedlings in the hole tray to a nursery, spreading organic fertilizer 4-7 cubic meters and urea 30 kg/mu in the nursery before transplanting, then deeply turning over, leveling the land, ridging and ridging, planting the furrows with the width of 80cm, planting 4 rows in each furrow, planting wide and narrow rows, planting the wide row spacing of 40cm, the narrow row spacing of 20cm, the plant spacing of 15-20cm and 50cm between furrows, fully watering and thoroughly watering the seedlings before transplanting, avoiding transplanting at noon and noon, starting transplanting at 2 o' clock after transplanting, watering the nursery after transplanting, completely surviving the seedlings after one month, topdressing from 5 to 8 mid-day days in 5 months, fertilizing 1 time every 10 days, and applying urea 2-4 kg/mu per time, after additional fertilization, a small amount of fertilizer is needed for multiple times, weeds are removed before the additional fertilization, and the fertilizer cannot be scattered on seedling leaves;

and step 9: pruning in the seedling stage, the seedlings are easy to have long branches and thorns, the seedlings are easy to scratch in the operation period, and the future inoculation operation is not facilitated, so that the branches and the thorns on the trunk are pruned in 8 months, and the branches and the thorns on the trunk are pruned once in the late 9 months;

step 10: preparing before inoculation, starting inoculation in the last 8 th day, selecting strains which are screened and have strong pathogenicity and wide distribution, preparing a PSA culture medium in advance, performing high-temperature and high-pressure sterilization at the environment of 121 ℃ for 25min, subpackaging on an ultraclean workbench by using sterile culture dishes, subpackaging each dish by a thin layer which cannot be too thick, inoculating the strains on the culture medium after solidification, culturing by using an incubator at the temperature of 25 ℃, beating hypha to a fungus cake by using a puncher after the hypha grows to be full of the culture dishes after 4 days, cutting filter paper into squares of 2cm multiplied by 2cm, performing high-pressure sterilization for later use, preparing a preservative film and cutting the filter paper into 5cm wide;

step 11: inoculating strong seedlings with the ground diameter exceeding 1cm, inoculating a fungus cake every 15cm from the ground height of hybrid seedlings of 80cm, namely inoculating the hybrid seedlings of 80cm, 95cm, 110cm, 125cm and 140cm once, punching the holes at the height until the fungus cake is thick to touch the xylem, placing the back of the prepared fungus cake on three layers of filter paper which is completely soaked by sterile water for moisture preservation, enabling the side with hyphae to face upwards, completely corresponding to the punched holes on the hybrid seedlings, slightly pressing the filter paper by hands for fixation, and completely winding the filter paper by a sealing film from bottom to top to keep the humidity;

step 12: the first investigation, the investigation is once after 3 weeks of inoculation, for can see the scab already extends to the single plant outside the plastic wrap, directly eliminate, cut off the miscellaneous seedling from the root, then collect and concentrate and burn;

step 13: and (4) second investigation: second investigation after 4 weeks of inoculation, unfastening a sealing film, removing filter paper, counting whether each inoculation point is attacked and the length of disease spots, calculating the morbidity and the morbidity degree of each individual plant, and grading the morbidity degree, wherein the morbidity = the inoculation point attacked/the total inoculation point x 100%, and the morbidity = the length of the total disease spots/5;

step 14: evaluating and eliminating the disease resistance of each plant, comprehensively judging the disease resistance of each plant by combining the disease incidence and the disease incidence degree, and determining the elimination amount of each combined plant, wherein the elimination force can be increased by large combinations with the number of the plants exceeding 1000 in the combination, and the single plants with the disease incidence rate exceeding 40% and the disease spot length larger than 1cm are eliminated; the number of single plants in the combination is less than 500, and single plants with the disease incidence rate of more than 80 percent and the length of disease spots of more than 2cm are eliminated;

step 15: burning eliminated plants, wherein the eliminated single plants are cut from the roots and then burned intensively so as not to be spread;

step 16: and (3) transferring the single plants, selecting the single plants sensitive and susceptible to the rot disease through inoculation identification, continuously grafting and culturing the remained single plants, and selecting the plumest buds within 10cm of the topmost end of the single plants in spring of the next year to graft the pear stocks with the row spacing of 0.5m multiplied by 2m of the planted plants.

The invention has the beneficial effects that: the method greatly improves the survival rate of the seedling plants through the steps of crossbreeding, seedling culture, single plant switching and the like, improves the rot resistance of the pear trees through breeding, improves the yield of the pear trees and increases economic benefits.

Detailed Description

The present invention will be further described with reference to the following examples.

Example 1

An efficient early-stage selective breeding method for pear trees to resist rot disease, which comprises the following steps:

step 1: selecting seeds, wherein the resistance of the filial generation to rot disease is influenced by the parent variety, the resistance of the parent variety has larger influence on the number of resistant plants of the filial generation, the number of disease-resistant plants in the filial generation of which parents are not the resistant variety is small, one of the parents or the parents is the disease-resistant variety, and more disease-resistant plants can be generated in the filial generation, so that the selected seeds are combined into one of the parents or the parents which is the disease-resistant variety, wherein the selected seeds are autumn pears and Chinese pears;

step 2: performing hybrid pollination, collecting buds in a big-balloon period in advance, stripping anthers, drying in an environment at 25 ℃ after collection, sealing in an environment at 4 ℃ after pollen scattering, temporarily storing until pollination is started in a big-balloon period of female parent buds, immediately bagging and isolating after pollination, and removing the isolation after pollination for 10 days;

and step 3: management of hybrid fruits, bagging fruits three weeks after pollination, spraying insecticide and bactericide once a week before bagging, carrying out insecticide and sterilization by adopting a solution of 800 times of thiophanate methyl 70% and a solution of 4000 times of abamectin 5%, soaking bag openings by using 500 times of thiophanate methyl and 1000 times of chlorpyrifos one night before bagging, firstly propping up the bag openings when bagging, propping up the bag bottoms, expanding the bag bodies, sealing the bag openings around fruit stalks after bagging fruits, carrying out pest control in the fruit growth period, strengthening fertilizer and water management, and paying attention to fruit-strengthening fertilizer for 5 months, wherein the fertilizer mainly comprises organic fertilizer, nitrogen fertilizer and potassium fertilizer, 30 kg of urea, 30 kg of potassium sulfate and 4000 kg of biogas slurry and water per mu is applied at this time;

and 4, step 4: harvesting the hybrid fruits, starting harvesting after the fruits are fully ripe, transporting the harvested fruits back to the room to allow the harvested fruits to emit field heat, removing fruit bags, and marking;

and 5: harvesting and refrigerating, wherein in order to fully after-ripen the seeds, the hybrid fruits are harvested and radiated, then precooled for 24 hours in the environment of about 20 ℃, and then stored in a refrigeration house at 0 ℃;

step 6: collecting seeds, refrigerating for 65 days, taking the fruits out of a refrigeration house, transversely cutting the fruits, taking the seeds out of ventricles by using tweezers, collecting the seeds by using seed bags, rubbing and washing the seeds at 25 ℃ warm water, drying the seeds at a ventilated position after washing, fully drying the seeds, marking and storing at room temperature;

and 7: seed stratification treatment: laminating seeds before and after the annual New year's day, putting the seeds in warm water at 25 ℃ to fully absorb water for 12 hours, preparing a preservation box, putting fully-absorbed filter paper at the bottom of the box, then uniformly and non-overlapping the seeds on the filter paper, checking once every 10 days, and after 80% of the seeds are exposed to the white, starting to prepare for sowing;

and 8: cultivating strong seedlings, sowing the seeds exposed to the white to a special hole tray of 11cm deep for forest trees, carrying out insecticidal and sterilization treatment on the matrix before sowing, transplanting when the seedling grows to 4 true leaves and the root is nearly 10cm, transplanting the seedling in the hole tray to a nursery, spreading and applying 7 cubic organic fertilizer and 30 kilograms of urea per mu in the nursery before transplanting, then deeply ploughing, leveling the land, ridging and ridging, wherein the width of each furrow is 80cm, 4 rows are planted in each furrow, the width of each row is wide and the width of each row is narrow, the row spacing of the wide rows is 40cm, the row spacing of the narrow rows is 20cm, the plant spacing is 15-20cm, and the space between the furrows is 50cm, the seedlings are thoroughly watered before being transplanted, transplanting is avoided at noon time, transplanting is started at 2 o' clock in the afternoon, the nursery garden after being transplanted is watered by sprinkling irrigation, the seedlings completely survive after one month, topdressing is carried out from 5 m.ten days to 8 m.ten days, fertilizer application is carried out for 1 time every 10 days, 3 kg/mu of urea is applied every time, and the fertilizer cannot;

and step 9: pruning in the seedling stage, the seedlings are easy to have long branches and thorns, the seedlings are easy to scratch in the operation period, and the future inoculation operation is not facilitated, so that the branches and the thorns on the trunk are pruned in 8 months, and the branches and the thorns on the trunk are pruned once in the late 9 months;

step 10: preparing before inoculation, starting inoculation in the last 8 th day, selecting strains which are screened and have strong pathogenicity and wide distribution, preparing a PSA culture medium in advance, performing high-temperature and high-pressure sterilization at the environment of 121 ℃ for 25min, subpackaging on an ultraclean workbench by using sterile culture dishes, subpackaging each dish by a thin layer which cannot be too thick, inoculating the strains on the culture medium after solidification, culturing by using an incubator at the temperature of 25 ℃, beating hypha to a fungus cake by using a puncher after the hypha grows to be full of the culture dishes after 4 days, cutting filter paper into squares of 2cm multiplied by 2cm, performing high-pressure sterilization for later use, preparing a preservative film and cutting the filter paper into 5cm wide;

step 11: inoculating strong seedlings with the ground diameter exceeding 1cm, inoculating a fungus cake every 15cm from the ground height of hybrid seedlings of 80cm, namely inoculating the hybrid seedlings of 80cm, 95cm, 110cm, 125cm and 140cm once, punching the holes at the height until the fungus cake is thick to touch the xylem, placing the back of the prepared fungus cake on three layers of filter paper which is completely soaked by sterile water for moisture preservation, enabling the side with hyphae to face upwards, completely corresponding to the punched holes on the hybrid seedlings, slightly pressing the filter paper by hands for fixation, and completely winding the filter paper by a sealing film from bottom to top to keep the humidity;

step 12: the first investigation, the investigation is once after 3 weeks of inoculation, for can see the scab already extends to the single plant outside the plastic wrap, directly eliminate, cut off the miscellaneous seedling from the root, then collect and concentrate and burn;

step 13: and (4) second investigation: second investigation after 4 weeks of inoculation, unfastening a sealing film, removing filter paper, counting whether each inoculation point is attacked and the length of disease spots, calculating the morbidity and the morbidity degree of each individual plant, and grading the morbidity degree, wherein the morbidity = the inoculation point attacked/the total inoculation point x 100%, and the morbidity = the length of the total disease spots/5;

step 14: evaluating and eliminating the disease resistance of each plant, comprehensively judging the disease resistance of each plant by combining the disease incidence and the disease incidence degree, and determining the elimination amount of each combined plant, wherein the elimination force can be increased by large combinations with the number of the plants exceeding 1000 in the combination, and the single plants with the disease incidence rate exceeding 40% and the disease spot length larger than 1cm are eliminated; the number of single plants in the combination is less than 500, and single plants with the disease incidence rate of more than 80 percent and the length of disease spots of more than 2cm are eliminated;

step 15: burning eliminated plants, wherein the eliminated single plants are cut from the roots and then burned intensively so as not to be spread;

step 16: and (3) transferring the single plants, selecting the single plants sensitive and susceptible to the rot disease through inoculation identification, continuously grafting and culturing the remained single plants, and selecting the plumest buds within 10cm of the topmost end of the single plants in spring of the next year to graft the pear stocks with the row spacing of 0.5m multiplied by 2m of the planted plants.

The method greatly improves the survival rate of the seedling plants through the steps of crossbreeding, seedling culture, single plant switching and the like, improves the rot resistance of the pear trees through breeding, improves the yield of the pear trees and increases economic benefits.

Claims (1)

1. An efficient rot disease-resistant early-stage selective breeding method for pear trees is characterized in that: the breeding method comprises the following steps:

step 1: selecting seeds, wherein the resistance of the filial generation to rot disease is influenced by the parent variety, the resistance of the parent variety has larger influence on the number of resistant plants of the filial generation, the number of the disease-resistant plants in the filial generation with parents not being the resistant variety is small, one of the parents or the parents is the disease-resistant variety, and more disease-resistant plants can be generated in the filial generation, so that the selected seeds are combined into one of the parents or the parents which is the disease-resistant variety, wherein the selected seeds are any two combinations of autumn pears, Chinese pears, white pears and Xinjiang pears;

step 2: performing hybrid pollination, collecting buds in a big-balloon period in advance, stripping anthers, drying in an environment at 25 ℃ after collection, sealing in an environment at 4 ℃ after pollen scattering, temporarily storing until pollination is started in a big-balloon period of female parent buds, immediately bagging and isolating after pollination, and removing the isolation after pollination for 10 days;

and step 3: management of hybrid fruits, bagging fruits three weeks after pollination, spraying insecticide and bactericide once a week before bagging, carrying out insecticide and sterilization by adopting a solution of 800 times of thiophanate methyl 70% and a solution of 4000 times of abamectin 5%, soaking bag openings by using 500 times of thiophanate methyl and 1000 times of chlorpyrifos one night before bagging, firstly propping up the bag openings when bagging, propping up the bag bottoms, expanding the bag bodies, sealing the bag openings around fruit stalks after bagging fruits, carrying out pest control in the fruit growth period, strengthening fertilizer and water management, and paying attention to fruit-strengthening fertilizer for 5 months, wherein the fertilizer mainly comprises organic fertilizer, nitrogen fertilizer and potassium fertilizer, 30 kg of urea, 30 kg of potassium sulfate and 4000 kg of biogas slurry and water per mu is applied at this time;

and 4, step 4: harvesting the hybrid fruits, starting harvesting after the fruits are fully ripe, transporting the harvested fruits back to the room to allow the harvested fruits to emit field heat, removing fruit bags, and marking;

and 5: harvesting and refrigerating, wherein in order to fully after-ripen the seeds, the hybrid fruits are harvested and radiated, then precooled for 24 hours in the environment of about 20 ℃, and then stored in a refrigeration house at 0-2 ℃;

step 6: collecting seeds, refrigerating for 60-80 days, taking the fruits out of a refrigerator, transversely cutting the fruits, taking the seeds out of ventricles by using tweezers, collecting the seeds by using seed bags, rubbing and washing the seeds at 25 ℃ in warm water, drying the seeds at a ventilated position after washing, fully drying the seeds, marking and storing at room temperature;

and 7: seed stratification treatment: laminating seeds before and after the annual New year's day, putting the seeds in warm water at 25 ℃ to fully absorb water for 12 hours, preparing a preservation box, putting fully-absorbed filter paper at the bottom of the box, then uniformly and non-overlapping the seeds on the filter paper, checking once every 10 days, and after 70-80% of the seeds are exposed to the white, starting to prepare for sowing;

and 8: cultivating strong seedlings, sowing the seeds after exposure to the white to a hole tray special for the forest with the depth of 11cm, carrying out insect killing and sterilization treatment on a matrix before sowing, starting transplanting when the seedlings grow to 4 real leaves and the roots are close to 10cm, transplanting the seedlings in the hole tray to a nursery, spreading organic fertilizer 4-7 cubic meters and urea 30 kg/mu in the nursery before transplanting, then deeply turning over, leveling the land, ridging and ridging, planting the furrows with the width of 80cm, planting 4 rows in each furrow, planting wide and narrow rows, planting the wide row spacing of 40cm, the narrow row spacing of 20cm, the plant spacing of 15-20cm and 50cm between furrows, fully watering and thoroughly watering the seedlings before transplanting, avoiding transplanting at noon and noon, starting transplanting at 2 o' clock after transplanting, watering the nursery after transplanting, completely surviving the seedlings after one month, topdressing from 5 to 8 mid-day days in 5 months, fertilizing 1 time every 10 days, and applying urea 2-4 kg/mu per time, after additional fertilization, a small amount of fertilizer is needed for multiple times, weeds are removed before the additional fertilization, and the fertilizer cannot be scattered on seedling leaves;

and step 9: pruning in the seedling stage, the seedlings are easy to have long branches and thorns, the seedlings are easy to scratch in the operation period, and the future inoculation operation is not facilitated, so that the branches and the thorns on the trunk are pruned in 8 months, and the branches and the thorns on the trunk are pruned once in the late 9 months;

step 10: preparing before inoculation, starting inoculation in the last 8 th day, selecting strains which are screened and have strong pathogenicity and wide distribution, preparing a PSA culture medium in advance, performing high-temperature and high-pressure sterilization at the environment of 121 ℃ for 25min, subpackaging on an ultraclean workbench by using sterile culture dishes, subpackaging each dish by a thin layer which cannot be too thick, inoculating the strains on the culture medium after solidification, culturing by using an incubator at the temperature of 25 ℃, beating hypha to a fungus cake by using a puncher after the hypha grows to be full of the culture dishes after 4 days, cutting filter paper into squares of 2cm multiplied by 2cm, performing high-pressure sterilization for later use, preparing a preservative film and cutting the filter paper into 5cm wide;

step 11: inoculating strong seedlings with the ground diameter exceeding 1cm, inoculating a fungus cake every 15cm from the ground height of hybrid seedlings of 80cm, namely inoculating the hybrid seedlings of 80cm, 95cm, 110cm, 125cm and 140cm once, punching the holes at the height until the fungus cake is thick to touch the xylem, placing the back of the prepared fungus cake on three layers of filter paper which is completely soaked by sterile water for moisture preservation, enabling the side with hyphae to face upwards, completely corresponding to the punched holes on the hybrid seedlings, slightly pressing the filter paper by hands for fixation, and completely winding the filter paper by a sealing film from bottom to top to keep the humidity;

step 12: the first investigation, the investigation is once after 3 weeks of inoculation, for can see the scab already extends to the single plant outside the plastic wrap, directly eliminate, cut off the miscellaneous seedling from the root, then collect and concentrate and burn;

step 13: and (4) second investigation: second investigation after 4 weeks of inoculation, unfastening a sealing film, removing filter paper, counting whether each inoculation point is attacked and the length of disease spots, calculating the morbidity and the morbidity degree of each individual plant, and grading the morbidity degree, wherein the morbidity = the inoculation point attacked/the total inoculation point x 100%, and the morbidity = the length of the total disease spots/5;

step 14: evaluating and eliminating the disease resistance of each plant, comprehensively judging the disease resistance of each plant by combining the disease incidence and the disease incidence degree, and determining the elimination amount of each combined plant, wherein the elimination force can be increased by large combinations with the number of the plants exceeding 1000 in the combination, and the single plants with the disease incidence rate exceeding 40% and the disease spot length larger than 1cm are eliminated; the number of single plants in the combination is less than 500, and single plants with the disease incidence rate of more than 80 percent and the length of disease spots of more than 2cm are eliminated;

step 15: burning eliminated plants, wherein the eliminated single plants are cut from the roots and then burned intensively so as not to be spread;

step 16: and (3) transferring the single plants, selecting the single plants sensitive and susceptible to the rot disease through inoculation identification, continuously grafting and culturing the remained single plants, and selecting the plumest buds within 10cm of the topmost end of the single plants in spring of the next year to graft the pear stocks with the row spacing of 0.5m multiplied by 2m of the planted plants.