Rice receptor kinase gene OsRLCK21, protein encoded by rice receptor kinase gene OsRLCK21and application

A technology of receptor kinase and gene encoding, applied in the field of genetic engineering, can solve the problems of easy loss of resistance and easy variation of rice varieties

Active Publication Date: 2020-01-17
INST OF CROP SCI CHINESE ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Xanthomonas oryzae pathogenic variants are easy to muta

Method used

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  • Rice receptor kinase gene OsRLCK21, protein encoded by rice receptor kinase gene OsRLCK21and application
  • Rice receptor kinase gene OsRLCK21, protein encoded by rice receptor kinase gene OsRLCK21and application
  • Rice receptor kinase gene OsRLCK21, protein encoded by rice receptor kinase gene OsRLCK21and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0046] Example 1. Method for knocking out rice receptor kinase gene OsRLCK21 based on Lb-CRISPR / Cpf1 system

[0047] 1. Sequence and analysis of rice receptor kinase gene OsRLCK21

[0048] The sequence of rice receptor kinase gene OsRLCK21 is shown in SEQ ID No.2. Sequence analysis shows that the gene includes 5 exons in total, which are the 1-3 (first exon), the 469-1154 (second exon), the 469-1154 (second exon) of the SEQ ID No. 2773-2808 (third exon), 3360-3688 (fourth exon), 3752-4429 (fifth exon).

[0049] In the present invention, the sequence on the second exon of the rice receptor kinase gene OsRLCK21 is used as the RLCK21-T1 target sequence of the method for knocking out the rice receptor kinase gene OsRLCK21 based on CRISPR / Cpf1 technology.

[0050] 2. Lb-CRISPR / Cpf1 vector primer design and construction of recombinant expression vector

[0051] 2.1 Selection of Lb-CRISPR / Cpf1 technology target sequence

[0052] The Lb-CRISPR / Cpf1 technology targets the sense str...

Embodiment 2

[0066] Example 2. Application of targeted knockout method in rice varieties based on Lb-CRISPR / Cpf1 technology

[0067] The recombinant Agrobacterium EH105-Lb-RLCK21-1 was used to infect the callus induced by mature embryos of the rice variety Nipponbare, and the obtained rice transformed plants were respectively named NIP-Lb-RLCK21-1; the specific methods of the experiment are as follows:

[0068] 1. Inoculate the recombinant Agrobacterium obtained in Example 1 into YEB liquid medium (containing 50 μg / ml kanamycin and 20 μg / ml rifampicin), and culture it with shaking at 28° C. and 200 rpm until the OD600 is 0.6-0.8 ; Centrifuge at 5000rpm and 4°C for 5min, and resuspend the bacterial pellet with AAM liquid medium (acetosyringone concentration: 200μM / L, pH 5.2) until the OD600 is 0.6-0.8.

[0069] 2. Remove the glumes from the mature seeds of the rice variety Nipponbare, soak them in 75% ethanol for 1 min, then sterilize them in NaClO solution (mixed with water 1:2, add 1 drop...

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Abstract

The invention relates to a rice receptor kinase gene OsRLCK21, a protein encoded by the rice receptor kinase gene OsRLCK21 and application. A nucleotide sequence of the rice receptor kinase gene OsRLCK21 is shown as SEQ ID NO.2, and a protein sequence encoded by the gene is shown as SEQ ID NO.1. The OsRLCK21 gene in rice is knocked out through CRISPR/Cpf1 technology in a fixed-point mode to perform a function study of the gene, a result shows that the rice knocked out of the OsRLCK21 gene has significantly improved resistance to Xanthomonas oryzae IV and GD1358, IV lesion length is shortened by 26.4%-48.8%, C5 lesion length is shortened by 43%-52.3%, rice gene mutations can be caused by inserting or deleting a 503 position to a 508 position in a sequence shown SEQ ID NO.2, and a trait of increased resistance to Xanthomonas oryzae is shown.

Description

technical field [0001] The invention relates to the technical field of genetic engineering, in particular to the rice receptor kinase gene OsRLCK21 and its encoded protein and application. Background technique [0002] Bacterial blight caused by Xanthomonas oryzae pv.oryzae is an important bacterial disease that restricts rice production. It can generally reduce rice production by about 20%-30%, and can reach 50% in severe cases. %, which is seriously harmful to the rice planting industry. Utilizing resistance genes and cultivating disease-resistant varieties is the most economical and effective measure to control rice bacterial blight. However, most of the 42 rice bacterial blight resistance genes / loci reported so far (http: / / www.shigen.nig.ac.jp / rice / oryzabase / gene / list) show a narrow resistance spectrum Or difficult to use, only genes such as Xa3, Xa4, Xa21 and Xa23 are widely used in production. [0003] There is a specific interaction between rice and Xanthomonas ory...

Claims

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Application Information

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IPC IPC(8): C12N15/54C12N9/12C12N15/84A01H5/00A01H6/46
CPCC12N9/12C12N15/8205C12N15/8281
Inventor 周永力李全林史晓荣
Owner INST OF CROP SCI CHINESE ACAD OF AGRI SCI
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