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CN-121575153-B - InDel molecular marker linked with wheat-wheatgrass introgression line powdery mildew resistance locus PmYZW-6A-1 and application thereof

CN121575153BCN 121575153 BCN121575153 BCN 121575153BCN-121575153-B

Abstract

The invention relates to the technical field of molecular biology, and provides an InDel molecular marker linked with a wheat-wheatgrass introgression system powdery mildew resistance locus PmYZW-6A-1 and application thereof. InDel molecular markers linked with powdery mildew resistance locus PmYZW-6A-1 of wheat-agropyron aegerita introgression line are positioned at positions 231-257 of a nucleotide sequence shown in SEQ ID NO.1, and resistance locus PmYZW-6A-1 is positioned at chromosome 442,022 and 338bp of common ice 3504 reference genome 6A. By the technical scheme, the problem that the traditional wheat breeding method cannot efficiently utilize genetic diversity and has low culture efficiency of broad-spectrum durable disease-resistant varieties is solved.

Inventors

  • LI LIHUI
  • LING HONGQING
  • ZHENG LI
  • ZHENG LIYAN

Assignees

  • 崖州湾国家实验室
  • 海南大学三亚南繁研究院

Dates

Publication Date
20260508
Application Date
20260128

Claims (7)

  1. 1. The InDel molecular marker for identifying the wheat powdery mildew resistance is characterized in that the nucleotide sequences of the InDel molecular marker are shown as SEQ ID NO.1 and SEQ ID NO.2, the nucleotide sequence shown as SEQ ID NO.1 is an insertion type, and the nucleotide sequence shown as SEQ ID NO.2 is a deletion type.
  2. 2. The application of the kit in detecting the powdery mildew resistance of wheat is characterized in that the kit comprises a primer pair; the primer pair is used for amplifying the InDel molecular marker according to claim 1, and comprises the following components: The nucleotide sequence of InDel-F is shown as SEQ ID NO. 3; The nucleotide sequence of InDel-R is shown as SEQ ID NO. 4; if a 289bp specific band is amplified by utilizing the primer pair, the wheat to be detected is identified as the genotype of the disease, If the primer pair is utilized to amplify a specific band of 262bp, the wheat to be detected is identified as the disease resistance genotype, If a 289bp specific band and a 262bp specific band are amplified by using the primer pair, the wheat to be detected is identified as heterozygous genotype.
  3. 3. The application of the primer pair in wheat genetic analysis is characterized in that the wheat genetic analysis is wheat powdery mildew resistance genetic diversity analysis, wheat powdery mildew resistance identification or powdery mildew resistance wheat auxiliary breeding, The primer pair is used for amplifying the InDel molecular marker according to claim 1, and comprises the following components: The nucleotide sequence of InDel-F is shown as SEQ ID NO. 3; The nucleotide sequence of InDel-R is shown as SEQ ID NO. 4; if a 289bp specific band is amplified by utilizing the primer pair, the wheat to be detected is identified as the genotype of the disease, If the primer pair is utilized to amplify a specific band of 262bp, the wheat to be detected is identified as the disease resistance genotype, If a 289bp specific band and a 262bp specific band are amplified by using the primer pair, the wheat to be detected is identified as heterozygous genotype.
  4. 4. The application of the kit in wheat genetic analysis is characterized in that the wheat genetic analysis is wheat powdery mildew resistance genetic diversity analysis, wheat powdery mildew resistance identification or powdery mildew resistance wheat auxiliary breeding; the kit comprises a primer pair; the primer pair is used for amplifying the InDel molecular marker according to claim 1, and comprises the following components: The nucleotide sequence of InDel-F is shown as SEQ ID NO. 3; The nucleotide sequence of InDel-R is shown as SEQ ID NO. 4; if a 289bp specific band is amplified by utilizing the primer pair, the wheat to be detected is identified as the genotype of the disease, If the primer pair is utilized to amplify a specific band of 262bp, the wheat to be detected is identified as the disease resistance genotype, If a 289bp specific band and a 262bp specific band are amplified by using the primer pair, the wheat to be detected is identified as heterozygous genotype.
  5. 5. A method for identifying wheat powdery mildew resistance is characterized by comprising the steps of taking genomic DNA of wheat to be detected as a template, carrying out PCR (polymerase chain reaction) amplification by using a primer pair, carrying out electrophoresis on an amplified product, and judging the wheat powdery mildew resistance according to an electrophoresis band type; the primer pair is used for amplifying the InDel molecular marker according to claim 1, and comprises the following components: The nucleotide sequence of InDel-F is shown as SEQ ID NO. 3; The nucleotide sequence of InDel-R is shown as SEQ ID NO. 4; The method for judging the powdery mildew resistance of the wheat according to the electrophoresis bands comprises the following steps: If only one 289bp specific band appears in the amplified product, judging that the genotype of the disease exists in the wheat to be detected; if only one 262bp specific band appears in the amplified product, judging that the disease-resistant genotype exists in the wheat to be detected; If the amplified product has bands of 289bp and 262bp, the heterozygous genotype of the wheat to be detected is judged.
  6. 6. A method for identifying wheat powdery mildew resistance according to claim 5, wherein the PCR amplification reaction system comprises 1 μL of template DNA, 1 μ L, ddH 2 O3 μL of 10 μM primer pair, 2× TAQ PCR MASTER Mix 5 μL of primer pair comprising InDel-F and InDel-R, and the PCR amplification reaction program comprises denaturation at 94 ℃ for 3min, denaturation at 94 ℃ for 30sec, annealing at 55 ℃ for 30sec, elongation at 72 ℃ for 30sec,35 cycles, and elongation at 72 ℃ for 5min.
  7. 7. The method for breeding powdery mildew resistant wheat is characterized by comprising the following steps: Taking genomic DNA of wheat to be detected as a template, carrying out PCR amplification by using a primer pair, carrying out electrophoresis on an amplified product, judging powdery mildew resistance of the wheat according to an electrophoresis band type, and if the amplified product only has one 262bp specific band, retaining the wheat to be detected for breeding; the primer pair is used for amplifying the InDel molecular marker according to claim 1, and comprises the following components: The nucleotide sequence of InDel-F is shown as SEQ ID NO. 3; The nucleotide sequence of InDel-R is shown as SEQ ID NO. 4.

Description

InDel molecular marker linked with wheat-wheatgrass introgression line powdery mildew resistance locus PmYZW-6A-1 and application thereof Technical Field The invention relates to the technical field of molecular biology, in particular to an InDel molecular marker linked with a wheat-wheatgrass introgression system powdery mildew resistance site PmYZW-6A-1 and application thereof. Background Wheat (triticum aestivum l.) is a gramineous wheat crop, belonging to the annual self-pollinating plant. Wheat is one of the major food crops worldwide, and the whole growth period is threatened by a plurality of diseases. Wheat powdery mildew is a fungal disease caused by Blumeria gramineae wheat specialization Blumeria agraminisf.sp.tritici (Bgt), and is one of the most destructive diseases on wheat. The development of disease-resistant breeding by using powdery mildew resistance genes is the most economical and effective prevention and control strategy. It has been shown that wheat resistance to powdery mildew can be divided into different levels according to genetic mechanisms, but relying on single resistance is prone to loss of resistance due to rapid evolution of pathogenic bacteria. Even in plants containing known powdery mildew resistance sites, more pathogenic powdery mildew isolates may still occur. Therefore, a highly effective disease-resistant barrier must be constructed by integrating multiple levels of resistance to prevent adaptive evolution of powdery mildew and break through resistance. Genetic diversity against race specificity and quantitative resistance exists widely in wheat, which provides a genetic basis for breeding varieties with broad-spectrum, durable disease resistance by a method combining phenotypes with molecular marker-assisted selection. Currently, the development and utilization of plant disease-resistant genes become the core direction of crop disease-resistant breeding, and the expansion of wheat powdery mildew-resistant gene libraries has important significance for the cultivation of novel disease-resistant varieties. Molecular breeding provides means and possibilities for accelerating conventional breeding processes. By understanding the functions of the parent, especially the major genes, the probability of cross-combining can be predicted, enabling the cultivation of a purported variety. The marker assisted selection can transfer the phenotype screening in the breeding process to early-stage accurate screening based on molecular markers, so that the breeding efficiency is improved. The further integration of the optimized marker assisted selection system and the high-throughput genotyping and breeding acceleration method can help to accelerate the breeding utilization of powdery mildew resistance genes, increase the genetic variation of wheat and provide support for the continuous prevention and control of powdery mildew. Disclosure of Invention The invention provides an InDel molecular marker linked with a wheat-wheatgrass introgression line powdery mildew resistance locus PmYZW-6A-1 and application thereof, and solves the problem of low culture efficiency of a broad-spectrum durable disease-resistant variety in a traditional wheat breeding method. The technical scheme of the invention is as follows: The invention provides an InDel molecular marker linked with a wheat-agropyron aegerita introgression system powdery mildew resistance site PmYZW-6A-1, wherein the InDel molecular marker is positioned at the 231 th-257 th positions of a nucleotide sequence shown in SEQ ID NO. 1. In the invention, inDel molecular markers are developed according to InDel variation of the whole genome sequencing result of the anti-sense mixing pool. As a further technical scheme, the resistance locus PmYZW-6A-1 is located at the chromosome 442,022,338bp of the reference genome 6A of pulmonic 3504. In the invention, the resistance locus PmYZW-6A-1 is a major powdery mildew resistance QTL locus positioned by analyzing a recombinant inbred line population of pulmonic 3504X She Kaola by utilizing a BSA-seq technology. The invention also provides a primer pair for amplifying the InDel molecular marker linked with the wheat-agropyron introgression system powdery mildew resistance locus PmYZW-6A-1, which comprises the following components: InDel-F5'-TTTTGGACGACAAGATGGTTATTTA-3', shown as SEQ ID NO. 3; InDel-R5'-ACTCCATTACAGTCTCACGTACCAG-3' is shown as SEQ ID NO. 4. The invention also provides a kit for detecting the powdery mildew resistance of wheat, which comprises the primer pair; if a 289bp specific band is amplified by utilizing the primer pair, the wheat to be detected is identified as the genotype of the disease, If the primer pair is utilized to amplify a specific band of 262bp, the wheat to be detected is identified as the disease resistance genotype, If a 289bp specific band and a 262bp specific band are amplified by using the primer pair, the wheat to be detected is identified as heterozygous genot