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Application of wheat puccinia striiformis westend.f.sp.tritici PSTG_13661 gene to puccinia striiformis westend.f.sp.tritici prevention and treatment and method for cultivating puccinia striiformis westend.f.sp.tritici-resistant wheat

A technology for wheat stripe rust and wheat stripe rust, which is applied in the fields of genetic engineering and crop molecular breeding, can solve the problems of difficulty in wheat-stripe rust genetic research, slow progress, and low number of key pathogenic genes, and achieves significant wheat-stripe rust. The effect of stripe rust resistance

Active Publication Date: 2018-05-08
SHANDONG AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
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AI Technical Summary

Problems solved by technology

[0004] Due to its biotrophic and obligate parasitic characteristics, genetic transformation of wheat stripe rust is difficult; and wheat itself is also difficult to carry out genetic transformation, so the genetic research of wheat-stripe rust interaction is very difficult and the progress is slow
At present, the number of infection or pathogenic key genes identified in wheat stripe rust is relatively small, and no genes have been effectively used in wheat disease resistance breeding

Method used

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  • Application of wheat puccinia striiformis westend.f.sp.tritici PSTG_13661 gene to puccinia striiformis westend.f.sp.tritici prevention and treatment and method for cultivating puccinia striiformis westend.f.sp.tritici-resistant wheat
  • Application of wheat puccinia striiformis westend.f.sp.tritici PSTG_13661 gene to puccinia striiformis westend.f.sp.tritici prevention and treatment and method for cultivating puccinia striiformis westend.f.sp.tritici-resistant wheat
  • Application of wheat puccinia striiformis westend.f.sp.tritici PSTG_13661 gene to puccinia striiformis westend.f.sp.tritici prevention and treatment and method for cultivating puccinia striiformis westend.f.sp.tritici-resistant wheat

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] (1) PCR amplification to obtain the PSTG_13661 gene fragment

[0043] Taking the complete genome sequence of wheat stripe rust physiological race PST-78 as a reference (GenBank accession number AJIL00000000.1 or BROAD download link ftp: / / ftp.broadinstitute.org / pub / annotation / fungi / puccinia / genomes / puccinia_striifo rmis_pst- 78 / ), using the sequence of hypothetical protein gene PSTG_13661 (GenBank: KNE92947.1) as a template to design PCR primers. The sequence of the forward primer CQM13661-F1 is shown in Seq ID No.3: 5`- CACAAGAAGGAATGGGTTTTACTC-3`; the sequence of the reverse primer CQM13661-R1 is shown in Seq ID No.4: 5`-CTCCAGGCCTAGCGAAGTCC-3`, CQM13661- F1 / R1 amplification amplifies the full-length coding region of the gene, and the length of the amplified fragment in the reference genome (physiological race PST-78) is 861 bp.

[0044] Compared with the gene structure PSTG_13661 annotated in the reference genome PST-78, the homologous gene amplified by CRY29 has a 7...

Embodiment 2

[0061] Embodiment 2 Anti-stripe rust resistance identification of wheat stripe rust

[0062] Propagation of fresh spores of wheat stripe rust: plant the susceptible variety Huixianhong in the greenhouse, inject the spore aqueous solution with a syringe during the one-leaf-one-heart stage, then spray water with a watering can and cover with a plastic film to keep it moist. Inoculation can be repeated 2-3 times to ensure full onset, with an interval of 1 week between each time. Preparation of the spore aqueous solution: Take the dried and frozen (-80°C) spores, suspend them with an appropriate amount of tap water until they are light orange, place them on a shaker at room temperature and shake them for 30 minutes (180rpm), and then inoculate them by injection. Sporadic disease can be seen about 20 days after inoculation, and a large number of leaves can be seen after about 30 days. A large number of fresh spores can be collected for inoculation identification of transgenic mater...

Embodiment 3

[0067] Example 3 Identification of PSTG_13661 Gene Expression in Stripe Rust-resistant Wheat

[0068] Twelve days after inoculation of wheat seedling leaves, the surface of wild-type CB037 leaves was covered with a large number of spore piles. About 100 mg of leaves of CB037 and transgenic materials were taken at different periods after wheat was inoculated with stripe rust, RNA was extracted by Trizol method, and cDNA was prepared for real-time fluorescent quantitative PCR. The primer sequences are CQM13661-F3 and CQM13661-R3, as shown in Seq ID No.10 and Seq ID No.11 in the sequence table. Taking the α-tubulin gene of wheat stripe rust as an internal reference (Huang Xueling et al., Journal of Agricultural Biotechnology, 2012, 20(2): 181-187), the primer TUBA-F / R sequences are shown in the sequence table Seq ID No.12, Seq ID No.13 shown.

[0069] The PCR system is: 5 μl 2×SYBR GreenMaster, 1 μl each of the forward and reverse primers (10 μM), 1 μl cDNA, 2 μl ddH2O, the tot...

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Abstract

The invention provides application of a puccinia striiformis westend.f.sp.tritici PSTG_13661 gene to puccinia striiformis westend.f.sp.tritici prevention and treatment. A sequence of the PSTG_13661 gene is shown as Seq ID No.5; the PSTG_13661 gene can effectively regulate and control the growth and the reproduction of puccinia striiformis westend.f.sp.tritici and is taken as a molecular target fortranscriptional regulation and control or a gene target for protein function inhibition, and the effect of inhibiting the growth and the reproduction of the puccinia striiformis westend.f.sp.triticiis achieved through silencing the PSTG_13661 gene. The wheat obtained according to a method for cultivating the puccinia striiformis westend.f.sp.tritici-resistant wheat by utilizing the gene has significant puccinia striiformis westend.f.sp.tritici resistance.

Description

technical field [0001] The invention belongs to the technical field of genetic engineering and crop molecular breeding, and in particular relates to the application of wheat stripe rust PSTG_13661 gene in the prevention and treatment of stripe rust and the cultivation method of stripe rust resistant wheat. Background technique [0002] Wheat is the second largest grain crop in my country, with an average annual sowing area of ​​about 360 million mu. The stability and improvement of wheat production are related to the national economy and people's livelihood. However, for many years, fungal diseases have posed a serious threat to wheat production, such as rust, powdery mildew, scab, sheath blight and other diseases, seriously affecting the yield and quality of wheat. Among them, wheat stripe rust, commonly known as "jaundice", is caused by obligate parasitic stripe rust (Puccinia striiformis Westend.f.sp.tritici). In my country, the incidence of wheat stripe rust is rampant...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/29C12N15/82A01H5/00A01H6/46
CPCC12N15/8282C07K14/415Y02A50/30
Inventor 吴佳洁黄德华张会飞刘强倪飞付道林
Owner SHANDONG AGRICULTURAL UNIVERSITY
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