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Long-chain non-coding RNA gene ALEX1 and application thereof to improvement of bacterial blight resistance of rice

A long-chain non-coding, gene technology, applied in the field of plant genetic engineering, can solve problems such as the application of disease resistance and defense response is still blank, achieve important theoretical significance and application value, and enhance the effect of resistance.

Active Publication Date: 2019-12-10
SUN YAT SEN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there are relatively few studies on lncRNA in plants, and the application of lncRNA to rice bacterial blight resistance and defense response is still blank so far.

Method used

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  • Long-chain non-coding RNA gene ALEX1 and application thereof to improvement of bacterial blight resistance of rice
  • Long-chain non-coding RNA gene ALEX1 and application thereof to improvement of bacterial blight resistance of rice
  • Long-chain non-coding RNA gene ALEX1 and application thereof to improvement of bacterial blight resistance of rice

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0037] Example 1 Screening of differentially expressed rice lncRNA after bacterial blight infection

[0038] In order to study the role of lncRNA in rice disease resistance response, the inventors performed strand-specific lncRNA sequencing on rice leaf samples infected with bacterial blight PXO99A for 0, 2h, 6h, 12h and 24h, and obtained 5.7×10 8 reads, of which 4.9×10 8 The reads matched the reference genome of rice, accounting for 85.26% of the total data obtained. After splicing and assembly, a total of more than 700,000 transcripts were obtained. After five basic screening procedures and coding potential screening, 48,727 rice lncRNA transcripts with high reliability were finally obtained. The cuffdiff software was used to quantitatively analyze the screened lncRNAs to obtain the FPKM information of the lncRNAs of each sample. The expression levels of 567 lncRNAs were significantly changed due to bacterial blight infection ( figure 1 ). To confirm the reliability of de...

Embodiment 2

[0039] Example 2 Acquisition and functional identification of plants activated by in situ expression of ALEX1

[0040] The 567 bacterial blight-responsive lncRNAs identified above are candidate genetic resources for regulating rice resistance to bacterial blight. In order to study whether the abnormal expression of these lncRNAs will affect the pathogenesis of rice bacterial blight, we searched the world's major rice mutant databases and identified a plant with activated lncRNA expression that exhibited obvious resistance to bacterial blight ( Figure 5 ). Since this lncRNA is highly specifically expressed only in leaves infected by bacterial blight, and is almost not expressed under normal physiological conditions ( Figure 4 ), so we named it ALEX1 (An LncRNA Expressed in Xoo-infected leaves), and the two activated expression lines of ALEX1 were named ALEX1ox-1 and ALEX1ox-2.

[0041] We have confirmed that ALEX1 is located on the antisense DNA strand of rice chromosome 8 ...

Embodiment 3

[0046] Example 3 In situ expression of ALEX1 activates plant exogenous methyl jasmonate treatment and determination of endogenous jasmonic acid content

[0047] In situ expression of wild-type Zhonghua 11 and ALEX1 activated rice seeds to germinate in water at 32°C for 3 days. Methyl jasmonate (Sigma Aldrich #392707) was diluted to the indicated concentrations in deionized water and used to treat 3-d-old seedlings as described above. The state of the roots was photographed after culturing for 6 days in a climate chamber at 27° C. and 70% humidity. Methyl jasmonate treatment experiments showed that JA signaling was enhanced in rice ALEX1-activated plants ( Figure 6 ).

[0048] JA, JA-Ile and OPDA were analyzed by liquid chromatography coupled to mass spectrometry (LC-MS / MS). Briefly, the tallest leaves of 3-week-old rice were homogenized in liquid nitrogen. A sample of a 200 mg powder sample of fresh tissue was added to 1,000 μL of 500:1 extraction buffer containing isopro...

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Abstract

The invention discloses a long-chain non-coding RNA gene ALEX1 and application thereof to improvement of bacterial blight resistance of rice. The long-chain non-coding RNA ALEX1 gene is cloned from the rice, the nucleotide sequence of the long-chain non-coding RNA ALEX1 gene is shown as SEQ ID NO:1, and a transcription product of the long-chain non-coding RNA ALEX1 gene is induced by xanthomonas oryzae pv and subjected to up-regulated expression. Through study and confirmation of biological functions of the ALEX1, by activating expression of the ALEX1, the content of jasmonic acid in the ricecan be increased, the resistance of the rice to the xanthomonas oryzae pv can be enhanced significantly, and it is shown that the long-chain non-coding RNA ALEX1 can positively regulate and control the resistance of the rice to the xanthomonas oryzae pv. A new strategy and genetic resources are provided for cultivating new rice disease-resistance varieties, especially new rice varieties resistingbacterial blight, and the very important theoretical significance and application value are achieved.

Description

technical field [0001] The invention relates to the technical field of plant genetic engineering, and more specifically, relates to the long-chain non-coding RNA gene ALEX1 and its application in improving rice bacterial blight resistance. Background technique [0002] Plant diseases seriously affect the yield and quality of crops, and the cultivation of disease-resistant plants is one of the major topics in the research of crop molecular breeding. Rice bacterial blight caused by the gram-negative bacterium Xanthomonas pathogenic strain Xoo (Xanthomonas oryzaepv.Oryzae) is one of the most common diseases in rice production. The vascular bundles proliferate and secrete a large amount of polysaccharides to block the vascular bundles, causing the leaves to wither and even die, seriously reducing the photosynthesis of rice, resulting in reduced or even no harvest of rice. [0003] At present, many genes related to rice bacterial blight resistance have been identified in the wor...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/113C12N15/11C12N15/82A01H5/00A01H6/46
CPCC12N15/113C12N15/8243C12N15/8251C12N15/8281C12N2310/11Y02A40/146
Inventor 于洋周燕飞张玉婵陈月琴
Owner SUN YAT SEN UNIV
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