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Paddy rice yield genotyping primer set based on KASP technology and use thereof

A primer combination and genotyping technology, which is applied in the fields of molecular biology and crop breeding, can solve the problems of large labor consumption, complicated operation, and the inability to accurately identify material gene haplotype combinations, and achieve low cost and easy operation. Simple, accurate and reliable results

Active Publication Date: 2017-07-11
广州固德生物技术有限公司
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, traditional linkage markers are mainly based on molecular markers linked to genes for genotype identification, which cannot be used to accurately identify favorable gene haplotype combinations for target traits of materials and carry out directional improved breeding, and the operation is complicated, the throughput is small, and the labor force high consumption

Method used

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  • Paddy rice yield genotyping primer set based on KASP technology and use thereof
  • Paddy rice yield genotyping primer set based on KASP technology and use thereof
  • Paddy rice yield genotyping primer set based on KASP technology and use thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] Example 1 Exploitation of rice important yield gene functional markers

[0029] 1. According to the literature Ashikari M, Sakakibara H, Lin S, et al. Cytokinin oxidase regulates rice grain production [J]. Science, 2005, 309(5735): 741-745.; Wang J, XuH, Li N, et al. Artificial selection of Gn1a plays an important role inimproving rice yields across different ecological regions[J].Rice,2015,8(1):37. Gn1a corresponds to the MSU gene number: LOC_Os01g10110, and the SNP at position 1605 in the coding region of the gene: T / G variation directly leads to gene function differences.

[0030] 2. According to the literature Fujita D, Trijatmiko K R, Tagle A G, et al. NAL1 allele from arice landrace greatly increases yield in modern indica cultivars [J]. Proceedings of the National Academy of Sciences, 2013, 110(51): 20431-20436 . NAL1 corresponds to the MSU gene number: LOC_Os04g52479, for the 698th SNP in the coding region of the gene: A / G variation directly leads to gene func...

Embodiment 2

[0045] Example 2 Polymorphism Screening of Rice Important Yield Gene Functional Markers

[0046] 1. Molecular markers for the functional sites of 6 important rice yield genes, for 12 rice varieties including African cultivated rice, indica rice (including Huanghuazhan, 93-11, etc., which are widely promoted in production), japonica rice, and common wild rice The quality materials were subjected to KASP reaction, and the marker typing was tested.

[0047] The system used in the PCR amplification reaction was 5 μl: template DNA 2.5 μl, primer mixture 0.07 μl, 2×KASP Master Mix 2.5 μl, the final concentrations of primers fam-F and primer hex-F were 12 μM, and the final concentration of primer R was 25 μM.

[0048] The conditions of the PCR amplification reaction are: pre-denaturation at 94°C for 15 minutes; the first step of amplification reaction: denaturation at 94°C for 20 seconds, gradient annealing at 61°C-55°C and extension for 60 seconds, 10 cycles, each cycle of annealing...

Embodiment 3

[0054] Example 3 Application of KASP marker in improving rice germplasm Zhenshan 97

[0055] 1. According to the genotyping results of rice germplasm yield in Table 4, Zhenshan 97 (ZS97B) only contains favorable variation in the yield gene Gn1a. Therefore, the germplasm materials with differences in the yield genes of Gn1a, NAL1, Ghd7.1 and Ghd8 were selected Nipponbare was crossed with ZS97B, and ZS97B was improved by directional selection.

[0056] 2. Use Gn1a-KASP to select Nipponbare and ZS97B true hybrid F1, backcross with ZS97B to harvest BC1F1, select NAL1, Ghd7.1 and Ghd8 gene heterozygous individual plants for planting, and backcross with ZS97B to harvest BC2F1.

[0057] 3. Select NAL1, Ghd7.1 and Ghd8 gene heterozygous individual plants for planting, and backcross with ZS97B to harvest BC3F1.

[0058] 4. Select NAL1, Ghd7.1 and Ghd8 gene heterozygous individual plants for planting, and backcross with ZS97B to harvest BC4F1.

[0059] 5. Plant a single heterozygous p...

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Abstract

The invention provides a paddy rice yield genotyping primer set developed based on a KASP technology. A paddy rice yield gene comprises Gn1a, NAL1, Ghd7, Hd1, Ghd7.1 and Ghd8. The sequences of the primer set are shown in the formulas of SEQ ID NO: 1 to 18. The set of the KASP functional markers provided by the invention and the primer set can quickly detect the combination mode of yield gene functional alleles in rice germplasm. The primer set has the advantages of accurate and reliable detection result, simple operation and low cost. The method can realize the detection of important yield functional genes of a rice breeding material, and provide scientific guidance for breeding and improvement of novel varieties of high yield rice.

Description

technical field [0001] The invention relates to the technical fields of molecular biology and crop breeding, in particular to a combination of primers for rice yield genotyping based on KASP technology and its application. Background technique [0002] In recent years, breeders have bred a large number of high-yielding rice varieties through the pedigree method. However, traditional hybridization and backcross selection often require large-scale populations and a large number of phenotypic investigations, and many phenotypic investigations are difficult, which greatly increases the workload and cost of breeding. With the cloning of important rice genes, the development of molecular markers of gene functional sites, the detection of gene combination types of target traits in breeding materials, and the introduction of corresponding good genes according to specific breeding improvement goals, it has become possible to breed new varieties. In addition, many traits in the breed...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/6858C12Q1/6895C12Q2600/13C12Q2600/156C12Q2531/113C12Q2563/107
Inventor 余四斌袁志阳孙文强熊银龚蓉张佳宁
Owner 广州固德生物技术有限公司
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