Application of OsAGO18 gene in improving rice stripe disease resistance of rice

A technology for leaf blight resistance, rice stripe, applied in the biological field

Active Publication Date: 2015-04-29
PEKING UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, this may not be a general phenomenon and the results may depend on the specific virus

Method used

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  • Application of OsAGO18 gene in improving rice stripe disease resistance of rice
  • Application of OsAGO18 gene in improving rice stripe disease resistance of rice
  • Application of OsAGO18 gene in improving rice stripe disease resistance of rice

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0061] Example 1. It was found that OsAGO18 gene can be induced and expressed by rice stripe virus

[0062] The OsAGO18 gene involved in this example is derived from rice (Oryza.sativa L.). The cDNA sequence of the OsAGO18 gene is shown in sequence 1 in the sequence table. Sequence 1 consists of 3267 nucleotides, of which 1-3267 is the coding sequence (ORF); sequence 1 encodes the protein shown in sequence 2 in the sequence listing, and sequence 2 consists of 1088 amino acid residues.

[0063] 1. Accumulation of OsAGO18 gene mRNA after rice stripe virus infection

[0064] Take 0.5 g each of the wild-type rice (Oryza. After medium grinding, total RNA was extracted according to Invitrogen's TRIzol Reagent instructions (Invitrogen Trizol Reagent, cat No. 15596-018). After measuring the concentration of total RNA, 10 μg of total RNA was taken, and the rice genomic DNA in the total RNA was digested according to the instructions of RQ1 DNase (Promega, product number: M610A). Dige...

Embodiment 2

[0100] Embodiment 2, OsAGO18 gene participates in rice anti-RSV defense response

[0101] 1. Obtaining homozygous mutants of ago18 rice

[0102] ago18 mutant seeds (NF6013) purchased in Tos17 database

[0103] (https: / / tos.nias.affrc.go.jp / ~miyao / pub / tos17 / ).

[0104] The T-DNA insert sequence is:

[0105] tos17_tail6 (LB): 5'-AGGTTGCAAGTTAGTTAAGA-3'.

[0106] Insertion site such as image 3 Shown in B.

[0107] Identification of homozygotes for ago18 mutants using methods provided in the database

[0108] (https: / / pc7080.abr.affrc.go.jp / cgibin / tos17 / ricegenome.cgi?action=getTarget&chr=7&pos=16895280&primer=y&version=7).

[0109] Using the homozygous cDNA of the rice ago18 mutant (NF6013) identified above as a template, semi-quantitative RT was performed using the 6013_LP / 6013_RP primer pair, tos17_tail6(LB) / 6013_RP primer pair, and OsAGO18-F / OsAGO18-R primer pair, respectively. -PCR reaction. At the same time, the wild-type rice (Oryza.sativa L.) variety Nipponbare wa...

Embodiment 3

[0160] Example 3, Enhanced disease resistance of transgenic rice overexpressing OsAGO18

[0161] 1. Construction of OsAGO18 gene plant expression vector

[0162] 1. Acquisition of rice cDNA template

[0163] According to the instructions, the total RNA of Nipponbare rice (Oryza sativa L.japonica.cv.Nipponbare) was extracted with TRIzol Reagent from Invitrogen, and reverse-transcribed with the company's SuperScript II reverse transcriptase. The primer used for reverse transcription was 16-nucleotide Oligod (T) primer, and the rice cDNA template obtained by reverse transcription was finally obtained.

[0164] 2. Acquisition of rice OsAGO18 gene

[0165] Using the rice cDNA obtained in step 1 as a template, PCR amplification was performed with the following primers.

[0166] AGO18cds-F: 5'-ATAATGGCGAGCCGAGGAGGAGGC-3' (positions 4-24 of this sequence are positions 1-21 of sequence 1, and ATA in the first three positions is a protective base);

[0167] AGO18cds-R: 5'-GACCTAGCAA...

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Abstract

The invention discloses application of an OsAGO18 gene in improving the rice stripe disease resistance of rice, and provides application of an OsAGO18 gene, encoding protein of the OsAGO18 gene, or a recombinant carrier containing the encoding gene, in regulating and controlling rice stripe disease resistance of plants. The amino acid sequence of the gene OsAGO18 protein is specifically shown in the sequence 2 in a sequence table. The experiment shows that the plant disease resistance in rice with over-expression of the OsAGO18 gene is improved; on the contrary, the disease resistance of an ago18 mutant is degraded, which means that the OsAGO18 protein encoded by the gene plays an important role in rice stripe disease resistance of rice.

Description

technical field [0001] The invention belongs to the field of biotechnology, and relates to the application of OsAGO18 gene in improving the resistance of rice to rice stripe blight. Background technique [0002] Rice stripe leaf blight is a viral disease caused by rice stripe virus (Rice Stripe Virus, RSV), which is transmitted by SBPH as a vector, commonly known as cancer on rice. Diseased plants often withered and booted or the ears were small and deformed. At the beginning of the rice seedling disease, intermittent yellow-green or yellow-white short stripes appear at the base of the heart leaf, and later the lesion enlarges and merges, expanding into yellow-green stripes parallel to the veins, and the stripes remain green; after jointing, the disease occurs on the sword. Yellow-green stripes appear on the lower part of the leaves, and all types of rice are not dead, but the heading is deformed and the fruit is few. [0003] Rice Stripe Virus (RSV) is a representative sp...

Claims

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Application Information

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IPC IPC(8): C12N15/82A01H5/00
Inventor 李毅戚益军吴建国杨志蕊郑立佳魏春红
Owner PEKING UNIV
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