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Application of puccinia striiformis PSTG_06025 gene to wheat stripe rust prevention and treatment and culture method of anti-puccinia striiformis 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 a small number of pathogenic key genes, difficulties in genetic research of wheat-stripe rust, difficulties in genetic transformation, etc. Growth and reproduction, significant effect of wheat stripe rust resistance

Active Publication Date: 2018-05-15
SHANDONG AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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 puccinia striiformis PSTG_06025 gene to wheat stripe rust prevention and treatment and culture method of anti-puccinia striiformis wheat
  • Application of puccinia striiformis PSTG_06025 gene to wheat stripe rust prevention and treatment and culture method of anti-puccinia striiformis wheat
  • Application of puccinia striiformis PSTG_06025 gene to wheat stripe rust prevention and treatment and culture method of anti-puccinia striiformis wheat

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0037] (1) PCR amplification to obtain the PSTG_06025 gene fragment

[0038] 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_striiformis_pst- 78 / ), PCR primers were designed using the coding sequence of the hypothetical protein gene PSTG_06025 (GenBank: KNF00609.1) as a template. The forward primer sequence CQM06025-F2 is shown in Seq ID No.3: 5`-caagcggccgcGACGGTCCTTTCGATTTTGAG-3`; the reverse primer sequence CQM06025-R2 is shown in Seq ID No.4: 5`-tcaggcgcgccATCTCCAGAATCGAATGTCCAG-3`. The length of the amplified segment is 369 bp. Using the sequence alignment of the amplified segments to compare the whole genome data of wheat stripe rust PST-78 ( https: / / genome.jgi.doe.gov / Pucst_PST78_ 1 / Pucst_PST78_1.home.html ) and the sequence of wheat stripe rust (Puccinia striiformisf.sp.tritic...

Embodiment 2

[0051] Embodiment 2 Anti-stripe rust resistance identification of wheat stripe rust

[0052] 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

[0057] Example 3 Identification of PSTG_06025 Gene Expression in Stripe Rust-resistant Wheat

[0058] Twelve days after inoculation of wheat seedling leaves, the surface of wild-type CB037 leaves was covered with a large number of spore piles. We took about 100 mg of leaves of CB037 and transgenic materials at different periods after wheat inoculation, extracted RNA by Trizol method, and prepared cDNA for real-time fluorescent quantitative PCR. The primer sequences are CQM06025-F3 and CQM06025-R3, and the specific sequences are shown in the sequence table SeqIDNo.7 and SeqIDNo.8. 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 sequence is shown in the sequence table SeqID No.9, Seq ID Shown in No.10.

[0059] 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 total system is ...

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Abstract

The invention provides application of a puccinia striiformis PSTG_06025 gene to wheat stripe rust prevention and treatment. The sequence of the PSTG_06025 gene is shown as Seq ID No.1. The PSTG_06025gene can be used for effectively regulating and controlling the growth and the breeding of the puccinia striiformis; the PSTG_06025 gene is used as a transcriptional control molecular target or a protein function inhibition gene target; the effect of inhibiting the growth and the breeding of the puccinia striiformis is achieved through silencing the PSTG_06025 gene. The wheat obtained by the method for culturing the anti-puccinia striiformis wheat by using the gene provided by invention has obvious puccinia striiformis 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_06025 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
CPCC07K14/415C12N15/8218C12N15/8282
Inventor 吴佳洁张会飞黄德华刘强倪飞付道林
Owner SHANDONG AGRICULTURAL UNIVERSITY
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