CN112216343B - Screening method of tomato spotted wilt virus resistant pepper plants - Google Patents
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Abstract
The invention discloses a screening method of tomato spotted wilt virus resistant pepper plants, which comprises the steps of obtaining phenotype data of a tested material for resisting TSWV; acquiring a key QTL locus resisting TSWV and a key SNP locus resisting TSWV; cloning and quantitatively expressing analysis are carried out on the screened candidate genes; on the basis of correlation analysis and linkage analysis of the whole genome of the capsicum anti-TSWV, the relation between the found polymorphic site and the anti-TSWV intensity is developed, and a PCR-based molecular marker is developed aiming at the obvious correlation site/QTL site; based on the determined molecular marker, selecting backbone inbred line resisting tomato spotted wilt virus, and selecting hybrid variety resisting tomato spotted wilt virus by three-line method. The screening method can provide theoretical and material support for the breeding of the capsicum disease-resistant molecules and provide more effective means for the breeding of new capsicum varieties, thereby screening out the characteristic hybridization combination which accords with the preparation of high-quality dry capsicum, and carrying out popularization and application, and has very remarkable economic and social benefits.
Description
Technical Field
The invention relates to the technical field of crop breeding protection, in particular to a screening method of tomato spotted wilt virus resistant pepper plants.
Background
The capsicum is a favorite vegetable crop in daily life, is widely cultivated, and is often easy to cause diseases and insect pests in the pepper planting process. Tomato spotted wilt virus (Tomato spotted wilt virus, TSWV) is an important plant virus that jeopardizes agricultural production, is a representative species of the genus tomato spotted wilt virus belonging to the family bunyaviridae, can infect more than 70 families of 1 thousand plants, is currently known to be the plant virus with the widest host range, is widely distributed throughout the world, is also the primary virus for pepper production in north america, europe, etc., is reported in succession in brazil, united states, italy, australia, korea, and causes great losses to pepper production, and can cause sterilization in severe cases. In recent years, there is also report of tomato spotted wilt virus found on Yunnan chilli in China. Tomato spotted wilt virus is a virus disease specially transmitted by thrips, frankliniella occidentalis is first discovered in China around 2000, and then the frankliniella occidentalis is popular in the whole country, and is seriously damaged in open fields of Guangdong, hainan, guangxi, yunnan and Guizhou, and in the greenhouse such as Shandong and Beijing in the North. With the epidemic spread of frankliniella occidentalis, tomato spotted wilt virus has a tendency to aggravate the damage to capsicum.
Because the toxin-transmitting insect frankliniella occidentalis has high fluidity and quick propagation, even through a method of improving cultivation measures and a cultivation structure and combining chemical control and biological control, the method has a very difficult effect of achieving obvious control, so that the method is a major problem which has plagued pepper production from discovery until now, and is deeply concerned by researchers at home and abroad. From the perspective of 'plant health management', the breeding of pepper virus disease resistant varieties is considered as the most urgent, most effective and most environmentally friendly way to go out, i.e. the disease resistance of hosts is one of the most effective ways to prevent diseases at present. Screening and evaluation of TSWV resistant germplasm resources of capsicum are sequentially developed at home and abroad, and research on disease resistance QTL positioning, germplasm innovation and the like based on molecular markers and related mapping groups (F2) is performed. However, due to strong research dispersibility, disease-resistant germplasm resources mainly come from foreign scientific research institutions, different QTL positioning groups, lack of common molecular markers, lack of fine positioning and gene cloning groups and other factors, the process of cultivating new species of high-resistance tomato spotted wilt virus capsicum is severely restricted and limited.
Disclosure of Invention
The invention aims at solving the technical problems in the background technology, provides a screening method, can accurately reflect the disease resistance of the tomato spotted wilt virus, and selects a characteristic hybridization composition which accords with the preparation of high-quality dry peppers to popularize and apply by identifying the disease resistance of each variety of pepper seedlings to the tomato spotted wilt virus, and particularly relates to a screening method of tomato spotted wilt virus resistant pepper plants.
In order to solve the technical problems, the invention adopts the following technical scheme: a screening method of tomato spotted wilt virus resistant pepper plants comprises the following steps:
(1) The method comprises the steps of performing phenotype identification on pepper seedlings of inter-species BA 3X YNCXML F2 groups and pepper seedlings of inbred lines or local species through single-point and repeated field tests respectively, and obtaining phenotype data of a tested material for resisting TSWV;
(2) Based on a high-density genetic linkage map constructed by inter-species BA 3X YNCXML F2 population, carrying out QTL positioning on the anti-TSWV of the capsicum by combining the phenotype data of the anti-TSWV of the tested material obtained in the step (1) to obtain key QTL sites of the anti-TSWV; meanwhile, based on SNP genotype data constructed by cross lines or local species of peppers, carrying out whole genome association analysis on the anti-TSWV of the peppers by combining the phenotype data of the anti-TSWV of the tested materials obtained in the step (1) to obtain key SNP loci of the anti-TSWV;
(3) Determining inter-section and intra-section gene distribution information on the genome sequence of the capsicum Zun la-1 on the basis of the key QTL locus and the key SNP locus obtained in the step (2), screening haplotypes or allelic variants with important application value by using a comparative genomics and bioinformatics analysis method, screening out key QTL and genes for determining TSWV resistance, and carrying out cloning and quantitative expression analysis on the screened candidate genes;
(4) On the basis of the whole genome association analysis and linkage analysis of the TSWV resistance of the capsicum, selecting 5-8 genome sequences in which SNP/QTL (single nucleotide polymorphism)/TSWV resistance are obviously associated, combining the selected gene functions with the detected QTL sites of the TSWV resistance, selecting 3-5 candidate genes, searching polymorphic sites of the genes by a PCR resequencing method, detecting the relation between the found polymorphic sites and the TSWV resistance by a candidate gene association analysis method, and developing PCR-based molecular markers aiming at the obvious association sites/QTL sites;
(5) Based on the molecular marker determined in the step (4), selecting backbone inbred line resisting tomato spotted wilt virus, and selecting hybrid variety resisting tomato spotted wilt virus by using three-line method.
In the step (4), the relation between the polymorphic site and the TSWV resistance is the disease of the pepper seedlings to be detected, the disease of the disease-resistant pepper seedlings to be detected needs to be investigated once every 10 days, the incidence A of the pepper seedlings to be detected, which is 2-3 times required to be investigated, resistant plants and inductive plants are counted, and the highest incidence investigation result is used as the disease condition of the pepper seedlings of the variety; morbidity a= (number of susceptible plants of the variety/total number of plants of the variety) ×100%.
Further, the screening method of the tomato spotted wilt virus resistant pepper plants comprises the steps of adopting a serological detection or a molecular biological detection method for disease investigation; when the serology detection is performed, collecting leaves of a pepper seedling plant with disease resistance to be detected, performing ELISA detection by using a tomato spotted wilt virus test strip or a tomato spotted wilt virus antibody, wherein if the leaf is positive, the leaf is a disease-sensitive plant, and otherwise, the leaf is a disease-resistant plant; and during the molecular biological detection, extracting total RNA of plants, and carrying out RT-PCR detection by using a tomato spotted wilt virus specific primer, wherein if positive, the positive is represented as a disease-resistant plant, and otherwise, the positive is represented as a disease-resistant plant.
Further, the screening method of the tomato spotted wilt virus resistant pepper plants comprises the steps of observing symptoms; symptoms of the infected plants at the time of symptom observation include: the system leaves are necrotic spots, shrunken and yellow, the plants are short and small, and the whole plant is necrotic in the later stage of the disease.
Furthermore, the screening method of the tomato spotted wilt virus resistant pepper plants disclosed by the invention has the advantages that the relation between the polymorphic locus and the TSWV resistance is divided into three types through the morbidity, wherein the morbidity A=0 is an immune variety, the morbidity a is more than 0 and less than or equal to 20 percent and is a disease resistant variety, and the morbidity a is more than 20 percent and less than or equal to 100 percent and is a disease resistant variety; finally, the disease-resistant variety is used as a molecular marker of PCR.
Compared with the prior art, the screening method of the tomato spotted wilt virus resistant pepper plants has the beneficial effects that: the method comprises the steps of obtaining QTL positioning and SNP loci by using phenotype data, searching the relation between the polymorphic loci and TSWV resistance on the basis of TSWV resistance whole genome association analysis and linkage analysis of the peppers, developing corresponding molecular markers for obvious association loci/QTL loci, breeding backbone inbred lines resistant to tomato spotted wilt viruses by the determined molecular markers, and finally breeding hybrid varieties resistant to tomato spotted wilt viruses by using a three-line method, thereby providing theory and material support for pepper disease-resistant molecular breeding and providing more effective means for breeding new pepper varieties, so that characteristic hybrid combinations which accord with preparation of high-quality dry peppers are screened, popularized and applied, and very remarkable economic and social benefits are achieved.
Detailed Description
The invention is further illustrated by the following specific examples, which should be understood to those skilled in the art that variations and modifications can be made without departing from the principles of the invention, and these should also be considered to be within the scope of the invention.
The reagents used in the examples of the present invention, if not specifically described, may be commercially available to carry out the present invention, and the technical schemes involved, if not specifically described, may be all carried out by techniques or conditions described in the literature in the art or according to the product specifications.
Example 1
A screening method of tomato spotted wilt virus resistant pepper plants comprises the following steps:
(1) Phenotypic identification of pepper against tomato spotted wilt virus: taking 297 parts of inter-species BA 3X YNCML F2 population and 399 parts of pepper inbred lines as research objects, and performing phenotype identification on the research objects through single-point and 2-year 2-point 4 repeated field experiments to obtain phenotype data of the test material for resisting TSWV; wherein the inbred line is a local Zunyi Capsicum annuum.
(2) QTL localization and genome-wide association analysis of pepper tomato spotted wilt virus: based on a high-density genetic linkage map constructed by inter-species BA 3X YNCXML F2 group, carrying out QTL positioning on the TSWV resistance of the capsicum by combining the phenotype data obtained in the step (1) to obtain a key QTL of the TSWV resistance; based on 399 parts of pepper inbred line or local SNP genotype data, carrying out genome-wide association analysis on the anti-TSWV of the peppers by combining the phenotype data obtained in the research content (1) to obtain key SNP loci of the anti-TSWV.
(3) Identification and analysis of key QTL/gene of tomato spotted wilt virus resistance of capsicum: based on the obtained key QTL locus and key SNP locus, the inter-section and intra-section gene distribution information is determined on the Zun la-1 genome sequence of the capsicum, haplotype or allele variation with important application value is screened by using comparative genomics and bioinformatics analysis strategies, the key QTL and gene for determining anti-TSWV are identified, and cloning and quantitative expression analysis are carried out on candidate genes.
(4) Digging tomato spotted wilt virus resistant genes of capsicum: on the basis of the whole genome association analysis and linkage analysis of the anti-TSWV of the capsicum, annotating genome sequences in which 5 SNP/QTL which are obviously associated with the anti-TSWV are located, selecting 3 candidate genes by combining the annotated gene functions with the detected QTL of the anti-TSWV, searching polymorphic sites of the genes by a PCR (polymerase chain reaction) resequencing method, detecting the relation between the polymorphic sites and the intensity of the anti-TSWV by the candidate gene association analysis method, and developing a PCR-based molecular marker aiming at the obvious association sites/QTL.
(5) Breeding and demonstration of hybrid new varieties resistant to tomato spotted wilt virus: based on the above molecular marker, selecting backbone inbred line of tomato spotted wilt virus, selecting new hybrid variety (combination) of tomato spotted wilt virus by three-line method, and selecting new hybrid variety of Zunyi pod pepper tomato spotted wilt virus for demonstration planting.
The relation between the polymorphic site and the TSWV resistance is the disease of the pepper seedlings to be detected, the disease of the disease-resistant pepper seedlings to be detected needs to be investigated every 10 days, the incidence A of the pepper seedlings to be detected against tomato spotted wilt viruses is calculated, the investigation is required to be carried out 2 times, the disease of the pepper seedlings is counted by dividing the resistant plants and the susceptible plants, and the highest incidence investigation result is used as the disease condition of the pepper seedlings of the variety; morbidity a= (number of susceptible plants of the variety/total number of plants of the variety) ×100%. The disease incidence is distinguished by the relation between the polymorphic site and the TSWV resistance, wherein the incidence A=0 is an immune variety, the incidence a is more than 0 and less than or equal to 20 percent and is a disease-resistant variety, and the incidence a is more than 20 percent and less than or equal to 100 percent and is a disease-resistant variety; finally, the disease-resistant variety is used as a molecular marker of PCR.
The disease investigation adopts a serological detection or molecular biological detection method; when the serology detection is performed, collecting leaves of a pepper seedling plant with disease resistance to be detected, performing ELISA detection by using a tomato spotted wilt virus test strip or a tomato spotted wilt virus antibody, wherein if the leaf is positive, the leaf is a disease-sensitive plant, and otherwise, the leaf is a disease-resistant plant; and during the molecular biological detection, extracting total RNA of plants, and carrying out RT-PCR detection by using a tomato spotted wilt virus specific primer, wherein if positive, the positive is represented as a disease-resistant plant, and otherwise, the positive is represented as a disease-resistant plant. While in symptomatic observation, symptoms of the infected plants include: the system leaves are necrotic spots, shrunken and yellow, the plants are short and small, and the whole plant is necrotic in the later stage of the disease.
Example 2
A screening method of tomato spotted wilt virus resistant pepper plants comprises the following steps:
(1) Phenotypic identification of pepper against tomato spotted wilt virus: taking 297 parts of inter-species BA 3X YNCML F2 population and 399 parts of pepper local species as research objects, and performing phenotype identification on the research objects through single-point and 2-year 2-point 4-repeated field experiments to obtain phenotype data of the test material for resisting TSWV; wherein the local species is Zunyi Zunjiao No. 4 capsicum in the Zunyi dynasty.
(2) QTL localization and genome-wide association analysis of pepper tomato spotted wilt virus: based on a high-density genetic linkage map constructed by inter-species BA 3X YNCXML F2 group, carrying out QTL positioning on the TSWV resistance of the capsicum by combining the phenotype data obtained in the step (1) to obtain a key QTL of the TSWV resistance; based on 399 parts of pepper inbred line or local SNP genotype data, carrying out genome-wide association analysis on the anti-TSWV of the peppers by combining the phenotype data obtained in the research content (1) to obtain key SNP loci of the anti-TSWV.
(3) Identification and analysis of key QTL/gene of tomato spotted wilt virus resistance of capsicum: based on the obtained key QTL locus and key SNP locus, the inter-section and intra-section gene distribution information is determined on the Zun la-1 genome sequence of the capsicum, haplotype or allele variation with important application value is screened by using comparative genomics and bioinformatics analysis strategies, the key QTL and gene for determining anti-TSWV are identified, and cloning and quantitative expression analysis are carried out on candidate genes.
(4) Digging tomato spotted wilt virus resistant genes of capsicum: on the basis of the whole genome association analysis and linkage analysis of the anti-TSWV of the capsicum, annotating genome sequences in which 6 SNP/QTL which are obviously associated with the anti-TSWV are located, selecting 4 candidate genes by combining the annotated gene functions with the detected QTL of the anti-TSWV, searching polymorphic sites of the genes by a PCR (polymerase chain reaction) resequencing method, detecting the relation between the polymorphic sites and the intensity of the anti-TSWV by the candidate gene association analysis method, and developing a PCR-based molecular marker aiming at the obvious association sites/QTL.
(5) Breeding and demonstration of hybrid new varieties resistant to tomato spotted wilt virus: based on the above molecular marker, selecting backbone inbred line of tomato spotted wilt virus, selecting new hybrid variety (combination) of tomato spotted wilt virus by three-line method, and selecting new hybrid variety of Zunyi pod pepper tomato spotted wilt virus for demonstration planting.
The relation between the polymorphic site and the TSWV resistance is the disease of the pepper seedlings to be detected, the disease of the disease-resistant pepper seedlings to be detected needs to be investigated every 10 days, the incidence A of the pepper seedlings to be detected against tomato spotted wilt viruses is calculated, the investigation is required for 3 times, the disease of the pepper seedlings is counted by dividing the resistant plants and the susceptible plants, and the highest incidence investigation result is used as the disease condition of the pepper seedlings of the variety; morbidity a= (number of susceptible plants of the variety/total number of plants of the variety) ×100%. The disease incidence is distinguished by the relation between the polymorphic site and the TSWV resistance, wherein the incidence A=0 is an immune variety, the incidence a is more than 0 and less than or equal to 20 percent and is a disease-resistant variety, and the incidence a is more than 20 percent and less than or equal to 100 percent and is a disease-resistant variety; finally, the disease-resistant variety is used as a molecular marker of PCR.
The disease investigation adopts a serological detection or molecular biological detection method; when the serology detection is performed, collecting leaves of a pepper seedling plant with disease resistance to be detected, performing ELISA detection by using a tomato spotted wilt virus test strip or a tomato spotted wilt virus antibody, wherein if the leaf is positive, the leaf is a disease-sensitive plant, and otherwise, the leaf is a disease-resistant plant; and during the molecular biological detection, extracting total RNA of plants, and carrying out RT-PCR detection by using a tomato spotted wilt virus specific primer, wherein if positive, the positive is represented as a disease-resistant plant, and otherwise, the positive is represented as a disease-resistant plant. While in symptomatic observation, symptoms of the infected plants include: the system leaves are necrotic spots, shrunken and yellow, the plants are short and small, and the whole plant is necrotic in the later stage of the disease.
Example 3
A screening method of tomato spotted wilt virus resistant pepper plants comprises the following steps:
(1) Phenotypic identification of pepper against tomato spotted wilt virus: taking 297 parts of inter-species BA 3X YNCML F2 population and 399 parts of pepper local species as research objects, and performing phenotype identification on the research objects through single-point and 2-year 2-point 4-repeated field experiments to obtain phenotype data of the test material for resisting TSWV; wherein the local species is Zunyi Zunjiao No. 5 of Zunyi Chilli in the morning.
(2) QTL localization and genome-wide association analysis of pepper tomato spotted wilt virus: based on a high-density genetic linkage map constructed by inter-species BA 3X YNCXML F2 group, carrying out QTL positioning on the TSWV resistance of the capsicum by combining the phenotype data obtained in the step (1) to obtain a key QTL of the TSWV resistance; based on 399 parts of pepper inbred line or local SNP genotype data, carrying out genome-wide association analysis on the anti-TSWV of the peppers by combining the phenotype data obtained in the research content (1) to obtain key SNP loci of the anti-TSWV.
(3) Identification and analysis of key QTL/gene of tomato spotted wilt virus resistance of capsicum: based on the obtained key QTL locus and key SNP locus, the inter-section and intra-section gene distribution information is determined on the Zun la-1 genome sequence of the capsicum, haplotype or allele variation with important application value is screened by using comparative genomics and bioinformatics analysis strategies, the key QTL and gene for determining anti-TSWV are identified, and cloning and quantitative expression analysis are carried out on candidate genes.
(4) Digging tomato spotted wilt virus resistant genes of capsicum: on the basis of the whole genome association analysis and linkage analysis of the anti-TSWV of the capsicum, annotating genome sequences in which 8 SNP/QTL which are obviously associated with the anti-TSWV are located, selecting 5 candidate genes by combining the annotated gene functions with the detected QTL of the anti-TSWV, searching polymorphic sites of the genes by a PCR (polymerase chain reaction) resequencing method, detecting the relation between the polymorphic sites and the intensity of the anti-TSWV by the candidate gene association analysis method, and developing a PCR-based molecular marker aiming at the obvious association sites/QTL.
(5) Breeding and demonstration of hybrid new varieties resistant to tomato spotted wilt virus: based on the above molecular marker, selecting backbone inbred line of tomato spotted wilt virus, selecting new hybrid variety (combination) of tomato spotted wilt virus by three-line method, and selecting new hybrid variety of Zunyi pod pepper tomato spotted wilt virus for demonstration planting.
The relation between the polymorphic site and the TSWV resistance is the disease of the pepper seedlings to be detected, the disease of the disease-resistant pepper seedlings to be detected needs to be investigated every 10 days, the incidence A of the pepper seedlings to be detected against tomato spotted wilt viruses is calculated, the investigation is required for 3 times, the disease of the pepper seedlings is counted by dividing the resistant plants and the susceptible plants, and the highest incidence investigation result is used as the disease condition of the pepper seedlings of the variety; morbidity a= (number of susceptible plants of the variety/total number of plants of the variety) ×100%. The disease incidence is distinguished by the relation between the polymorphic site and the TSWV resistance, wherein the incidence A=0 is an immune variety, the incidence a is more than 0 and less than or equal to 20 percent and is a disease-resistant variety, and the incidence a is more than 20 percent and less than or equal to 100 percent and is a disease-resistant variety; finally, the disease-resistant variety is used as a molecular marker of PCR.
The disease investigation adopts a serological detection or molecular biological detection method; when the serology detection is performed, collecting leaves of a pepper seedling plant with disease resistance to be detected, performing ELISA detection by using a tomato spotted wilt virus test strip or a tomato spotted wilt virus antibody, wherein if the leaf is positive, the leaf is a disease-sensitive plant, and otherwise, the leaf is a disease-resistant plant; and during the molecular biological detection, extracting total RNA of plants, and carrying out RT-PCR detection by using a tomato spotted wilt virus specific primer, wherein if positive, the positive is represented as a disease-resistant plant, and otherwise, the positive is represented as a disease-resistant plant. While in symptomatic observation, symptoms of the infected plants include: the system leaves are necrotic spots, shrunken and yellow, the plants are short and small, and the whole plant is necrotic in the later stage of the disease.
By adopting the screening method, the QTL/gene of the pepper for resisting the tomato spotted wilt virus is determined by constructing a phenotype database, and the influence of candidate genes on the pepper for resisting the tomato spotted wilt virus is utilized, so that a functional PCR marker which can be directly applied to molecular breeding research is developed; determining haplotypes (regulatory elements and genes) or allelic variations within a target chromosome segment that have important application value; finally, the three-line method is used for breeding new hybrid varieties (combinations) resisting the tomato spotted wilt virus, so that the new hybrid varieties resisting the tomato spotted wilt virus of Zunyi pod peppers are screened for demonstration planting. Thus, the characteristic hybridization combination which accords with the high-quality dry pepper planted locally can be selected for popularization and application, and more remarkable economic benefit and social benefit can be generated.
Claims (3)
1. The screening method of the tomato spotted wilt virus resistant pepper plants is characterized by comprising the following steps of:
(1) The method comprises the steps of performing phenotype identification on pepper seedlings of inter-species BA 3X YNCXML F2 groups and pepper seedlings of inbred lines or local species through single-point and repeated field tests respectively, and obtaining phenotype data of a tested material for resisting TSWV;
(2) Based on a high-density genetic linkage map constructed by inter-species BA 3X YNCXML F2 population, carrying out QTL positioning on the anti-TSWV of the capsicum by combining the phenotype data of the anti-TSWV of the tested material obtained in the step (1) to obtain key QTL sites of the anti-TSWV; meanwhile, based on SNP genotype data constructed by cross lines or local species of peppers, carrying out whole genome association analysis on the anti-TSWV of the peppers by combining the phenotype data of the anti-TSWV of the tested materials obtained in the step (1) to obtain key SNP loci of the anti-TSWV;
(3) Determining inter-section and intra-section gene distribution information on the genome sequence of the capsicum Zun la-1 on the basis of the key QTL locus and the key SNP locus obtained in the step (2), screening haplotypes or allelic variants with important application value by using a comparative genomics and bioinformatics analysis method, screening out key QTL and genes for determining TSWV resistance, and carrying out cloning and quantitative expression analysis on the screened candidate genes;
(4) On the basis of the whole genome association analysis and linkage analysis of the TSWV resistance of the capsicum, selecting 5-8 genome sequences in which SNP/QTL (single nucleotide polymorphism)/TSWV resistance are obviously associated, combining the selected gene functions with the detected QTL sites of the TSWV resistance, selecting 3-5 candidate genes, searching polymorphic sites of the genes by a PCR resequencing method, detecting the relation between the found polymorphic sites and the TSWV resistance by a candidate gene association analysis method, and developing PCR-based molecular markers aiming at the obvious association sites/QTL sites; the relation between the polymorphic site and the TSWV resistance is the disease of the pepper seedlings to be detected, the disease of the pepper seedlings to be detected needs to be investigated once every 10 days, the incidence A of the pepper seedlings to be detected against tomato spotted wilt viruses is calculated, the investigation is carried out 2-3 times in total, the resistance and the susceptibility plants are separated, and the highest incidence investigation result is used as the disease condition of the pepper seedlings of the variety to be counted; morbidity a= (number of susceptible plants of the variety/total number of plants of the variety) ×100%; the relation between the polymorphic site and the TSWV resistance is divided into three types by the morbidity, wherein the morbidity A=0 is an immune variety, the morbidity A is more than 0 and less than or equal to 20 percent and is a disease-resistant variety, and the morbidity A is more than 20 percent and less than or equal to 100 percent and is a disease-resistant variety; finally, using the disease-resistant variety as a PCR molecular marker;
(5) Based on the molecular marker determined in the step (4), selecting backbone inbred line resisting tomato spotted wilt virus, and selecting hybrid variety resisting tomato spotted wilt virus by using three-line method.
2. The method for screening pepper plants resistant to tomato spotted wilt virus according to claim 1, characterized by: the disease investigation adopts a serological detection or molecular biological detection method; when the serology detection is performed, collecting leaves of a pepper seedling plant with disease resistance to be detected, performing ELISA detection by using a tomato spotted wilt virus test strip or a tomato spotted wilt virus antibody, wherein if the leaf is positive, the leaf is a disease-sensitive plant, and otherwise, the leaf is a disease-resistant plant; and during the molecular biological detection, extracting total RNA of plants, and carrying out RT-PCR detection by using a tomato spotted wilt virus specific primer, wherein if positive, the positive is represented as a disease-resistant plant, and otherwise, the positive is represented as a disease-resistant plant.
3. The method for screening pepper plants resistant to tomato spotted wilt virus according to claim 2, characterized by: the disease investigation method also comprises symptom observation; symptoms of the infected plants at the time of symptom observation include: the system leaves are necrotic spots, shrunken and yellow, the plants are short and small, and the whole plant is necrotic in the later stage of the disease.
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Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4221419A (en) * | 1977-02-19 | 1980-09-09 | Keith Riley | Gripping devices |
| CN105907755A (en) * | 2016-06-10 | 2016-08-31 | 中国农业科学院蔬菜花卉研究所 | Molecular marker closely linked with resistance of hot peppers to cucumber mosaic virus and application of molecular marker |
| CN109182575A (en) * | 2018-09-10 | 2019-01-11 | 广东省农业科学院作物研究所 | A kind of method of the anti-southern rust inbred line of sweet corn of molecular marking supplementary breeding |
| CN110777197A (en) * | 2019-08-02 | 2020-02-11 | 中国农业科学院棉花研究所 | Major QTL method for rapidly identifying cotton-related traits through compound BSA-seq |
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Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4221419A (en) * | 1977-02-19 | 1980-09-09 | Keith Riley | Gripping devices |
| CN105907755A (en) * | 2016-06-10 | 2016-08-31 | 中国农业科学院蔬菜花卉研究所 | Molecular marker closely linked with resistance of hot peppers to cucumber mosaic virus and application of molecular marker |
| CN109182575A (en) * | 2018-09-10 | 2019-01-11 | 广东省农业科学院作物研究所 | A kind of method of the anti-southern rust inbred line of sweet corn of molecular marking supplementary breeding |
| CN110777197A (en) * | 2019-08-02 | 2020-02-11 | 中国农业科学院棉花研究所 | Major QTL method for rapidly identifying cotton-related traits through compound BSA-seq |
Non-Patent Citations (1)
| Title |
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| 辣椒中可用于辅助选择的分子标记及其研究进展;华明艳等;长江蔬菜(第06期);第14-17页 * |
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