96 results about "Tomato yellow leaf curl virus" patented technology

The invention discloses a method for preventing and treating tomato yellow leaf curl virus diseases. The method includes cultivating tomatoes in sunlight greenhouses in large rows and small rows; transplanting abutilon theophrasti 15-20 days before the tomatoes are planted in fields; planting one row of abutilon theophrasti at the planting distances of 20cm along the inner side of the root of a north wall of each greenhouse; planting a row of abutilon theophrasti at the planting distances of 2.0m between each two adjacent small field planting rows; pulling out the abutilon theophrasti among the small field planting rows after fruits of fifth panicles of the tomatoes are set. The row spacing of the large rows of the tomatoes planted in the sunlight greenhouses is 80cm, and the row spacing of the small rows of the tomatoes planted in the sunlight greenhouses is 60cm. The leaf age of the abutilon theophrasti is 4 leaves and 1 core when the abutilon theophrasti is transplanted. The method has the advantages that the incidence of the tomato yellow leaf virus diseases can be effectively reduced, and the incidence of tomato yellow leaf virus disease infected tomato varieties can be within the range of 10% under the control generally.

The invention belongs to the technical field of plant molecular marker technology, and particularly relates to a double-SNP marker of a gene Ty-1 related to resistance to tomato yellow leaf curl virus disease, cDNA sequences of disease-resistant gene Ty-1 and disease-infected gene (namely allele gene) ty-1 and positions thereof on a whole genome can be obtained from the reported WO2012125025 document. Through sequence comparison, the disease-infected and disease-resistant specific primers NL2 and NL3 can be designed respectively by selecting two multielement SNP-locus GT/CG and GTG/ACA, and are mixed to form a NL2-3 double-SNP marker primer. The three types of Ty-1 and ty-1 can be detected by the NL2-3SNP marker through once PCR and gel electrophoresis display.

The invention discloses a dot enzyme-linked immunosorbent assay (ELISA) method for detecting bemisia tabaci carrying a tomato yellow leaf curl virus and application of the method. Field bemisia tabaci is collected, pretreated and grinded, monoclonal antibody of the tomato yellow leaf curl virus is used, the dot-ELISA method which is used for detecting virus transmission medium bemisia tabaci is established, a quick kit is developed and researched, and the tomato yellow leaf curl virus which is carried by the bemisia tabaci is detected. By the aid of the method, the virus carrying rate of the field bemisia tabaci can be surveyed in a large scale, and the field tomato yellow leaf curl disease outbreak trend is predicted. The method is quick, efficient, sensitive, special, high in flux and simple to operate. The tomato yellow leaf curl disease can be diagnosed quickly, predicted and controlled scientifically.

The invention discloses a molecular marker auxiliary selection method for tomato yellow leaf curl viral disease and knot nematode. The method is characterized by comprising the following steps of: (1) extracting DNA from a tomato material F1 and a selfed separation offspring F2 individual; (2) performing multiple PCR amplification on the tomato material F1 and the selfed separation offspring F2; and (3) performing resistance evaluation on the breeding separation generation material according to the detection result. The multiple PCR amplification method can amplify Ty-1 genes and Mi genes in the same reaction system at the same time, so the method greatly simplifies experiment operation and experiment expense, is suitable for identifying and analyzing a large amount of sample material, greatly shortens the time and reduces the workload.

The invention discloses a quick, simple and effective tomato yellow leaf curl virus resistance determination method and belongs to the field of plant protection. Traditionally, the determination of the tomato yellow leaf curl virus resistance is performed by a Bemisia tabaci (Gennadius) inoculation determination method, while the Bemisia tabaci (Gennadius) inoculation determination method has difficulty in providing uniform and consistent inoculation pressure due to the influences (such as the number of the inoculated Bemisia tabaci (Gennadius), virus rate and feeding habit) of the Bemisia tabaci (Gennadius) and at the same time, suffers interference from other Bemisia tabaci (Gennadius) transmitted viruses, so the accuracy of the determination result is low. The agrobacterium rhizogenes injection inoculation method well solves the problem. The resistance of a tomato material to the tomato yellow leaf curl viruses can be determined by using constructed infectious clones and directly inoculating the tomato yellow leaf curl viruses by injection in stalks. The method avoids culturing Bemisia tabaci (Gennadius), ensures simple operation, high efficiency, high repeatability and high controllability, provides a relatively consistent pressure, obtains accurate result and is a simple and effective inoculation determination method.

The invention relates to an InDel molecular marker based on a TYLCV (tomato yellow leaf curl virus) resistance gene Ty-3. The nucleotide sequence of the molecular marker is represented as SEQ ID No.1. The invention further relates to a primer pair for amplifying the InDel molecular marker based on the TYLCV resistance gene Ty-3. Sequences of the primer pair are as follows: a forward primer sequence: 5'-TGCAGGAACAGAATGATAGAAAA-3', and a reverse primer sequence: 5'-GCTCAGCATCACCTGAGACA-3'. The molecular marker is totally based on the TYLCV resistance gene Ty-3. According to the developed InDel molecular marker based on the TYLCV resistance gene Ty-3, the marker is totally based on the resistance gene Ty-3, is completely linked with the resistance gene Ty-3 and is a codominant molecular marker. When the marker is applied to molecular marker-assisted selection of the TYLCV resistance gene Ty-3, the accuracy of selection can be greatly improved, the breeding cycle can be shortened, and accordingly, the breeding process is accelerated.

The invention provides a method for quickly and synchronously detecting two virus components, including the tomato yellow leaf curl virus and the accompanying China tomato yellow leaf curl virus satellite, on tomatoes, and belongs to the field of plant protection. According to the nucleotide sequence of the tomato yellow leaf curl virus and the nucleotide sequence of the China tomato yellow leaf curl virus satellite, two pairs of primers are designed, namely TYmF/TYmR and TycnbF/TycnbR, wherein the TYmF/TYmR combination is used for detecting the tomato yellow leaf curl virus, and the TycnbF/TycnbR combination is sued for detecting the China tomato yellow leaf curl virus satellite. After total DNAs are extracted from a sample, only one time of PCR is needed to achieve detection of the two components including the tomato yellow leaf curl virus and the accompanying China tomato yellow leaf curl virus satellite. The method solves the problems existing in traditional biology, serology, electron microscopy observation and other methods. Compared with the traditional method that a virus is detected through one-time PCR, the detection method reduces cost, saves time and is a special, sensitive, economical and convenient method.

The invention discloses a hybridoma cell strain capable of secreting a tomato yellow leaf curl virus (TYLCV) monoclonal antibody and application of the monoclonal antibody. A coat protein gene of a tomato yellow leaf curl virus separator (TYLCV-SH2) is cloned; the coat protein of the virus is expressed by a prokaryotic expression system; BALB/c mice are immunized by using expression and purification protein as antigen; a hybridoma cell strain D10 capable of performing passage stably and secreting the TYLCV monoclonal antibody is obtained through cell fusion, screening and cloning; and the collection number is CGMCC No.5538. The D10 monoclonal antibody ascites indirect ELISA valence reaches more than 10<-6>; and the antibody type and subclass are IgG1 and kappa chains. A dot-ELISA detection method for detecting the TYLCV of the tomatoes is established by the D10 monoclonal antibody; and when the leaf suffering from the disease is diluted according to the ratio of 1:320 (w/v and g/mL), the virus can still be detected. By the dot-ELISA and Tissue-blot ELISA method, the TYLCV of tomato samples in the field can be detected accurately, specifically and sensitively. Due to establishment of a preparation method for the TYLCV monoclonal antibody and a detection method, technological and material support is provided for diagnosis, prediction and scientific prevention and control of the tomato virus disease.

The invention provides a fast, simple and effective method for identifying resistance to tomato yellow leaf curl virus, belonging to the field of plant protection. The tomato yellow leaf curl virus is a virus spread by bemisa babaci, the multi-sampling bemisa babaci inoculation identification method is applied to the traditional virus resistance identification; however, as affected by the bemisa babaci (such as inoculation quantity, virus-carrying rate and chow characteristic of the bemisa babaci) the bemisa babaci inoculation identification method is very difficult to provide uniform inoculation pressure and at the same time is interfered by other virus spread by the bemisa babaci, which leads to poor accuracy and repeatability of the identification result. However, the toothpick inoculation method well solves these problems, uses the constructed infective cloning, inoculates the tomato yellow leaf curl virus by puncturing the phloem of a tomato by a toothpick and identifies the resistance of the tomato material to the tomato yellow leaf curl virus. The method of the invention has the characteristics of no need to raise the bemisa babaci, simple operation, high efficiency, good repeatability, good controllability, accurate identification result and capability of providing relatively uniform inoculation pressure, thus being a simple and effective inoculation identification method.

The invention provides a method for rapidly identifying two similar viruses (a tomato yellow leaf curl virus and a papaya leaf curl China virus) on a tomato, belonging to the field of plant protection. The method comprises the following steps: according to the nucleotide sequences of the tomato yellow leaf curl virus and the papaya leaf curl China virus, designing three primers, i.e. PA_F362, TY F833 and PATY_R, wherein the PA _ F362 and the PATY_ R are combined to detect the papaya leaf curl China virus, and the TY _ F833 and PATY _ R are combined to detect the tomato yellow leaf curl virus;and extracting total DNA of samples, and carrying out PCR only once, thus realizing identifying the tomato yellow leaf curl virus and the papaya leaf curl China virus. The method of the invention overcomes the problems of the traditional methods in biology, serology, electron microscope observation and the like; and compared with the method for detecting a virus by PCP once, the method of the invention reduces cost and saves time, thus being a specific, sensitive, economical and convenient detection method.

The invention discloses a kit for rapidly detecting tomato yellow leaf curl virus and application thereof. A pair of specific primers is designed according to the nucleotide sequence of a dominant strain of tomato yellow leaf curl virus in our country, and also an extraction buffer and a washing buffer for processing a sample are designed. By using the kit for rapid detection on tomato yellow leaf curl virus, a virus-inflected tomato sample does not need extraction of DNA, only a small amount of a disease leaf (about 10 mg) is needed, and the disease leaf only needs once PCR reaction after being processed by the extraction buffer and the washing buffer, so that identification on tomato yellow leaf curl virus is realized. Experiments prove that the method has relatively high sensitivity and the sensitivity reaches 1E4. Also, the method is capable of performing virus containing identification on virus-transmission vectors Q-biotype bemisia tabaci and B-biotype bemisia tabaci of tomato yellow leaf curl virus. Compared with traditionally biology and serology, the method is a simple, convenient, rapid, economic and sensitive method for detecting tomato yellow leaf curl virus.

The invention provides a method for fast inoculating a tomato yellow leaf curl virus and belongs to the field of plant protection. The tomato yellow leaf curl virus is fast inoculated by a stem injection method by using built infectious clone PTYj01. The tomato yellow leaf curl virus is frequently inoculated by the conventional bemisia tabaci gennadius inoculation method in which the bemisia tabaci gennadius needs to be specially fed, and the inoculation efficiency is influenced by the virus transmitting efficiency of the desmone insect and is interfered by the virus transmitting of other bemisia tabaci. The method effectively overcomes the defects of the conventional bemisia tabaci gennadius inoculation method, and has the advantages of no need of feeding the bemisia tabaci gennadius, simple operation, high efficiency, high repeatability, and high controllability, and is a simple and convenient and effective inoculation method.

The invention discloses a method and a kit for synchronously detecting four viruses inducing a tomato yellow leaf curl disease and belongs to the field of plant protection. The four viruses include tomato yellow leaf curl virus, Taiwan tomato leaf curl virus, Guangdong tomato yellow leaf curl virus and Guangdong tomato leaf curl virus. Four pairs of specific primers are designed; total DNA of a sample which is to be detected is extracted; the extracted DNA is taken as a template and the four pairs of specific primers are taken as primers to perform PCR (polymerase chain reaction) in the same system; the PCR product is detected; and according to the fragment size of the PCR product, the virus which invades the sample can be identified. The method and the kit are quick, simple, convenient, accurate and low in cost, can be used for synchronously detecting the four viruses which induce the tomato yellow leaf curl disease, and can be applied to the actual production to quickly, accurately and effectively detect the plant disease sample.

The invention discloses a method of detecting tomato yellow leaf curl virus based on RPA (recombinase polymerase amplification). The invention provides a primer pair for detecting tomato yellow leaf curl virus; the primer pair includes a primer pair (a) composed of two single-stranded DNA molecules shown by sequence 1 and sequence 2 in a sequence table, and a primer (b) as functional as the primer pair (a) and composed of two single-stranded DNA molecules shown by sequences formed by subjecting the sequence 1 and sequence 2 in the sequence table to substitution and/or deletion and/or addition of one or more nucleotides. The RPA primers herein can effectively amplify target genes, the specificity reaches 100%, and sensitivity reaches 6*105 c/mu L; the primers show no cross-reaction with common tomato viruses. The method acts as an available quick detection method for screening plantlets of nurseries, and is significant to preventing spreading of tomato yellow leaf curve virus.

The present invention relates to the field of tomato plants having an intense phenotype and Tomato Yellow Leaf Curl Virus (TYLCV) resistance.

The invention discloses a preventing and treating method of the tomato yellow leaf curl virus. Tomatoes are cultured in a solar greenhouse according to size, wherein the large line spacing is 80 cm, and the small line spacing is 60 cm; gathering inducing plants are transplanted 15-20 days before tomatoes are planted, wherein a line of gathering inducing plants are planted between two lines of planted tomatoes, and the row spacing is 25 cm; substrate seedling trays containing carbonized rice husks are adopted for culturing seedlings of the gathering inducing plants, carbonized rice husks are additionally applied in planting pits, and seedling tray dipping and root irrigating are conducted through liquid thiamethoxam; before the gathering inducing plants are transplanted, a vent hole of thesolar greenhouse is closed by a 40-mesh sophocarpidine pest-killing protection net. On the premise of greatly reducing the chemical pesticide consumption and achieving main crop non-contact chemical prevention and treatment, the morbidity of the tomato yellow leaf curl virus can be effectively reduced, and the morbidity of the tomato yellow leaf curl virus of the infected tomato species can generally be controlled to be 4% or lower.

The invention discloses an SSR (simple sequence repeat) marker for tomato yellow leaf curl disease resisting character co-segregation and application thereof. The SSR marker includes primers: upstream primer T5-F: 5'-CAATCAATCGATGCACAAAACACC-3'; downstream primer T5-R: 5'-GACTGACTGCATTGGATTTGGCTT-3'. The marker can be amplified in CLN32120a-23 into a 580bp strip, can be amplified in Money marker into a 640bp strip and can be amplified in F1 into the above two strips, marking results are substantially consistent to that of field identification, it is proved that the marker can distinguish anti-disease materials, disease-sensitive materials and hybrid anti-disease materials and is a co-dominant marker in tight linkage with tomato yellow leaf curl disease virus resisting gene ty-5. Molecular marker detection is 90% matching with field detection in terms of detection results. The marker can be used in PCR (polymerase chain reaction) quick detection for tomato yellow leaf curl virus resisting ty-5 gene, and certain theoretical and practical basis is provided for the application of ty-5 molecular marker assisted selective breeding.

The invention discloses a method for cultivating a high-TYLCV (tomato yellow leaf curl virus)-resistant tomatoes. The method comprises the following steps: selecting fragments of AV1 gene, AC1 gene and AC3 gene in a TYLCV genome for gomphosis to obtain polygenes chimera AV1-AC1-AV3, and constructing an interference vector RNAi of a palindromic sequence; carrying out transient expression of dsRNA on tomato cotyledon by virtue of agrobacterium-mediated transformation to obtain a transgenic tomato for transforming the polygenes chimera AV1-AC1-AV3; and if the transgenic tomato shows TYLCV resistance after system infection of the TYLC, is not observed with obvious symptoms and has high-TYLCV resistance, screening out the high-TYLCV-resistant tomato. By utilizing the method, the high-TYLCV-resistant tomato strains can be quickly screened out and cultivated in 2 months, the cultivation period is greatly shortened, and a technical support is provided for preventing and curing TYLCV.