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SNP marker co-separated from maize gray leaf spot resistant main effect QTL-qRgls1 and application of SNP marker

A gray spot disease and co-segregation technology, applied in DNA/RNA fragments, recombinant DNA technology, microbial measurement/inspection, etc., can solve the problems of unfriendly environment, high cost, low throughput, etc., to save manpower and material resources and Time, short time-consuming, and the effect of reducing detection costs

Active Publication Date: 2019-04-12
YUAN LONGPING HIGH TECH AGRI CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] At present, the use of molecular markers such as SSR to screen disease resistance QTLs has disadvantages such as low throughput, high cost, and unfriendly environment, and it is difficult to be widely used in commercial breeding.

Method used

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  • SNP marker co-separated from maize gray leaf spot resistant main effect QTL-qRgls1 and application of SNP marker
  • SNP marker co-separated from maize gray leaf spot resistant main effect QTL-qRgls1 and application of SNP marker
  • SNP marker co-separated from maize gray leaf spot resistant main effect QTL-qRgls1 and application of SNP marker

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0038] Example 1 Acquisition of SNP markers associated with gray spot disease

[0039] It is known that the QTL locus qRgls1 related to resistance to maize gray spot disease is located on chromosome 8 of maize. The disease gene sequence is shown in SEQ ID NO: 2; Utilize the sequence alignment software DNAMAN to compare the nucleotide sequences of SEQ ID NO: 1 and 2, select 7 single nucleotide polymorphisms (SNP, Single Nucleotide Polymorphism) sites: HB1-1, HB1-2, HB1-3, HB1-4, HB1-5, HB1-6, HB1-7, and extract SNP sites HB1-1, HB1-2, HB1- 3. The nucleotide sequences of at least 50 bp on both sides of HB1-4, HB1-5, HB1-6, and HB1-7, respectively, such as SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6. Shown in SEQ ID NO: 7, SEQ ID NO: 8, and SEQ ID NO: 9. The principles for screening SNP sites are as follows: ① There are no other SNP sites in the vicinity of the SNP site; ② There are no InDel sites within 50 bp of the selected SNP site; ③ The sequences around the sele...

Embodiment 2

[0040] Embodiment 2KASP primer design and verification

[0041] The obtained SNP sites HB1-1, HB1-2, HB1-3, HB1-4, HB1-5, HB1-6, HB1-7 and the sequences on both sides such as SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9 (the 51st base n of each sequence is a SNP site, SEQ ID NO: 3-9 The corresponding base n is c / t, c / t, g / t, a / c, c / t, g / a, a / g (the disease resistance corresponding to each base is disease resistance / susceptibility ). Submit to Aiji Analysis (Shanghai) Technology Co., Ltd. for KASP primer design, the specific primer sequence is as follows:

[0042]

[0043] Among them, the 5' ends of Primer_FAM and Primer_HEX are respectively added with tag sequences, the tag sequence of FAM is 5'-GAAGGTGACCAAGTTCATGCT-3', and the tag sequence of HEX is 5'-GAAGGTCGGAGTCAACGGATT-3'.

[0044] The above-mentioned KASP primers were used to detect the corresponding SNP sites, and the application effect of the KASP primers was ver...

Embodiment 3

[0045] Example 3 Use of SNP markers for assisted selection in the backcross improvement process of gray spot disease

[0046] Using the disease-resistant material Y32 containing the gray spot disease resistance gene as the donor parent, and the material LA2061 susceptible to gray spot disease as the recurrent parent, the disease resistance gene was introduced into the recurrent parent LA2061 by backcrossing, and the KASP primer combination HB1- 3 and the SNP high-throughput detection platform produced by LGC for backcross generation BC 2 f 1 individual plants were tested. Specifically: take fresh leaves of each individual plant in the field, a total of 182 samples, and use the hotshot method to quickly extract leaf DNA. Use the LGCOKTOPURE DNA automatic extraction workstation to transfer the extracted DNA from the 96-well plate to the 384-well plate, use the LGCIntelliqube SNP high-throughput detection platform to divide and mix the PCR reaction system into the array tape, a...

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Abstract

The invention provides an SNP marker co-separated from maize gray leaf spot resistant main effect QTL-qRgls1 and an application of the SNP marker. The SNP marker is selected from at least one of markers SNP1-SNP3, and the nucleotide sequences of the markers are respectively shown in SEQID NO:5-7, wherein the 51st base of each marker is polymorphism sites. The invention further provides a KASP primer for detecting the SNP marker. The SNP marker and the KASP primer thereof provided by the invention can efficiently identify materials being in resistance to gray leaf spot in maize in low cost, andhigh-flux detection of resistance genes resistant to gray leaf spot in maize can be realized.

Description

technical field [0001] The invention belongs to the fields of molecular biology and plant molecular breeding, and in particular relates to a SNP marker co-separated with the main effect QTL-qRgls1 of maize gray spot resistance and its application. Background technique [0002] Corn is the most important food, feed, industrial raw material and energy crop in the world today. Gray spot is a fungal disease caused by Cecrospora zeae-maydis Tehon & Daniels, also known as Cecrospora zeae-maydis Tehon & Daniels. Gray spot disease was first discovered in the United States in the 1920s, and now it has become one of the main diseases that endanger corn production, especially in Southwest my country. Long-term production practice has proved that breeding and popularizing disease-resistant varieties is one of the most effective ways to control corn gray spot. [0003] For maize disease resistance breeding, the identification of disease resistance of breeding materials is very importan...

Claims

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Application Information

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IPC IPC(8): C12Q1/6895C12N15/11
CPCC12Q1/6895C12Q2600/13C12Q2600/156
Inventor 邹继军卢东林徐明良马传禹郑小明阮祥经李晓鹏谈存梅马丽
Owner YUAN LONGPING HIGH TECH AGRI CO LTD
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