EST-STS marking primer 1 with differential rye1RS chromosome, screening method and use

A technology of EST-STS and labeled primers, which is applied in the direction of biochemical equipment and methods, measurement/inspection of microorganisms, DNA/RNA fragments, etc., can solve problems such as inability to identify and distinguish, and achieve broadened application range and binding capacity Strong, loose experimental conditions

Inactive Publication Date: 2008-04-23
INST OF GENETICS & DEVELOPMENTAL BIOLOGY CHINESE ACAD OF SCI
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Problems solved by technology

However, these markers are amplified for the whole genome of rye and are not specific for 1R or 1RS chromosomes, making it impossible to associate 1R chromosomes with 2R chromosomes in rye. ～ Chromosome 7R separates
Koebner et al. synthesized the rye 1R chromosome-specific primer NOR-R1 based on the sequence difference between rye and wheat rRNA intergenic regions, which can identify the 1RS chromosome, but the primer NOR-R1 is used for recombinant materials that do not contain the nucleolus tissue region on the 1R chromosome. Will not be identified (reference: Koebner R.M.D. 1995. Generation of PCR-based markers for the detection of rye chromatin in wheat background. Theor Appl Genet, 90: 740-745)

Method used

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  • EST-STS marking primer 1 with differential rye1RS chromosome, screening method and use
  • EST-STS marking primer 1 with differential rye1RS chromosome, screening method and use
  • EST-STS marking primer 1 with differential rye1RS chromosome, screening method and use

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Embodiment 1

[0029] Example 1. Design and screening of primers

[0030] 1. Obtaining EST sequence: According to the wheat expressed sequence tag (EST) database published on the website of the United States Department of Agriculture (http: / / wheat.pw.usda.gov / GG2 / index.shtml), search and locate the first part of wheat homology The ESTs and sequences of different physical segments of the group, select 35 ESTs with strong conservation and base number greater than 300bp for download. Figure 1 is the base sequence of one of EST-BE637935.

[0031] 2. Design and synthesis of primers: use PrimerPremier 5.0 software to design primers for the EST sequence downloaded in step 1, and a total of 35 pairs of primers were designed. The conditions for primer design are annealing temperature 50-60℃, primer length 18-22bp, GC% content (guanine and Cytosine in the total number of bases) 40-60%, and the expected amplification product is 300-600bp . The designed primers were synthesized by Shanghai United Gene Co., ...

Embodiment 2

[0039] Example 2. The detection application of the labeled primer of the present invention in 1RS translocation material

[0040] Using the specific marker primer STS of the rye 1RS chromosome screened in Example 1 WE126 , Expand Xiaoyan 6 and "German White Tablet ” Part of the offspring material of S.cereale L.cv germanwhite hybridization, the reaction conditions and the amplification procedure are the same as step 4. After the amplification reaction, the amplified products are detected, and the detection method is the same as that in step 4. Among these materials, 4 materials amplified a specific band with a size of 850 bp to confirm that they contained the rye 1RS chromosome, and the other 7 materials did not amplify this corresponding specific band, and then it was determined that they did not contain rye 1RS. Chromosomes, as shown in Figure 5. This result is consistent with the identification result of genome in situ hybridization.

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Abstract

The present invention provides one kind of specific EST-STS marking primer-1 of rye 1RS chromosome. The marking primer-1 is designed with the EST sequence, BE637935, of the first partial wheat homologous group as the template, and has the sequences including STSWE126F of 5'-TCAAGCACGCATTTCAACTC-3' and STSWE126R of 5'-ACAGATGTCCAAAGCCCAAC-3'. The primer STSWE126 has one amplified 850 bp size specific segment located in rye 1RS chromosome. The marking primer-1 of the present invention expands the use range of wheat EST, provides one new tool for deep research of distant hybrid material for common wheat. The marking primer-1 may be applied in identifying rye 1RS chromosome simply and fast by means of common PCR technology.

Description

Technical field [0001] The invention relates to a specific EST-STS marker primer 1 for rye 1RS chromosome 1, a screening method and application, and belongs to the field of gene molecular marker research. Background technique [0002] Wheat is one of the most important food crops in the world. It is often threatened by various diseases in its production, resulting in a decline in yield or quality. Rye, a closely related species of wheat, has many excellent characteristics such as disease resistance, high yield, and barren tolerance. Studies have shown that there are many resistance genes on the rye 1RS chromosome, such as powdery mildew resistance, stripe rust resistance, leaf rust resistance, rod rust resistance, and wheat aphid resistance, as well as genes that increase yield and adaptability. Through distance hybridization and chromosome manipulation techniques, these beneficial genes on the rye 1RS chromosome can be introduced into common wheat. [0003] After the rye 1RS chr...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12N15/11C12N15/10
Inventor 王春梅安调过高强
Owner INST OF GENETICS & DEVELOPMENTAL BIOLOGY CHINESE ACAD OF SCI
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