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Tomato non-coding RNA gene LelncRNA1 and application thereof

A non-coding, genetic technology, applied in the field of plant genetic engineering, can solve the problems of limited important functions and unreported anti-viral disease functions, and achieve the effect of improving disease resistance

Inactive Publication Date: 2016-01-27
JIANGSU ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, it is known that the important functions of lncRNA in plants are limited to flowering regulation (Swiezewskietal., 2009; Heoetal., 2011; Sunetal., 2013), rice photosensitive genic male sterility (Dingetal., 2012; Zhouetal., 2012), Arabidopsis thaliana Fusarium wilt resistance (Zhuetal., 2014), corn anther development (Maetal., 2008), etc., the function of antiviral disease has not been reported

Method used

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  • Tomato non-coding RNA gene LelncRNA1 and application thereof
  • Tomato non-coding RNA gene LelncRNA1 and application thereof
  • Tomato non-coding RNA gene LelncRNA1 and application thereof

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Experimental program
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Effect test

Embodiment 1

[0025] A tomato variety CLN2777A resistant to TYLCV was selected (Liu Tingli, Yang Mali, Liu Xiaoshuang, Pei Yanfei, Zhao Huijuan, Yang Yuwen, Chen Tianzi, Zhang Baolong. Functional analysis of a tomato gene LeFLS2 induced by TYLCV-induced up-regulation. Journal of Jiangsu Agricultural Science, 2014, 30(2) :376-380), when the tomato grows to the 3-4 leaf stage, the tomato leaves are quick-frozen in liquid nitrogen, ground in a mortar, and the total RNA is extracted with the RNA extraction kit (DP419) of Beijing Tiangen Company, and the total RNA is identified by electrophoresis 质量;根据番茄RNA-Seq数据(TianziChen,YuandaLv,TongmingZhao,NanLi,YuwenYang,WenguiYu,XinHe,TingliLiu,BaolongZhang.Comparativetranscriptomeprofilingofaresistantvs.susceptibletomato(Solanumlycopersicum)cultivarinresponsetoinfectionbytomatoyellowleafcurlvirus.PLoSOne,2013,8(11):e80816),发现一个EST During the process of TYLCV infecting tomato, the expression was up-regulated, and the 5'RACE primers were designed according ...

Embodiment 2

[0029] The primers JAGN920F and JAGN921R designed in Example 1 were used to carry out quantitative RT-PCR analysis to inoculate the expression of 0d, 6d, and 12d tomato leaf LelncRNA1 gene after inoculating TYLCV. The results showed that after inoculating TYLCV, as the inoculation time became longer, the expression of LelncRNA1 gene Gradually enhanced, the expression level of 12dLelncRNA1 gene was about 2 times of 0d after inoculation ( image 3), indicating that the expression of LelncRNA1 gene is related to TYLCV infection.

Embodiment 3

[0031] According to the full-length sequence SEQIDNO.1 of the tomato LelncRNA1 gene, primers JAGN2225FCGCGGATCCAGCAGCAAATCCAGCGACCCGAG and JAGN2226RCGGGGTACCGTTGAATAATATTTAAGTTCACATCAGCA were designed to introduce restriction endonuclease sites BamHI and KpnI respectively, and the pMD19-T:LelncRNA1 plasmid obtained in step 2 was used as a template for PCR amplification Finally, LelncRNA1 was cloned into the binary expression vector pCAMBIA2301 (snapgene), sequenced and identified to ensure that the reading frame of the coding region in the expression vector was correct, and the vector with correct sequencing was named: pCAMBIA2301:LelncRNA1.

[0032] The expression vector pCAMBIA2301:LelncRNA1 was transferred into Agrobacterium strain EHA105 (Second Laboratory Company), and transferred into Ben's Tobacco (Nicotianabenthamiana), after PCR and RT-PCR verification of the obtained transgenic plants ( Figure 4 ), the positive transgenic T1 generation plants grown to the 3-4 leaf s...

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Abstract

The invention relates to tomato non-coding RNA gene LelncRNA1 and application thereof, and belongs to the field of plant genetic engineering. The cDNA sequence SEQ ID NO.1 of the gene is firstly reported in tomato filed, mRNA expression analysis shows that the gene shows up-regulated expression under induction effect of TYLCV, an obtained transgenic tobacco plant subjected to PCR and RT-PCR verification is inoculated with TYLCV for disease resistance evaluation, and results show that the transgenic plant shows obvious resistance to TYLCV compared with a contrast group. Therefore, LelncRNA1 gene can be introduced into plants as a target gene for improving virus disease resistance of plants.

Description

1. Technical field [0001] The invention belongs to the field of plant genetic engineering. Provided is a tomato non-coding RNA gene LelncRNA1 and its application for improving plant disease resistance through plant genetic engineering technology. 2. Background technology [0002] Studies have shown that 90% of human genomic DNA is transcribable (Wilhelmetal., 2008), and only 1–2% can encode proteins (Birney et al., 2007). Those RNA molecules that do not have the function of encoding proteins are called non-coding RNAs (ncRNAs ), wherein the non-coding RNA with a length greater than 200 nucleotides is called long non-coding RNA (Longnon-coding RNA, lncRNA) (Ghildiya and Zamore, 2009). lncRNA plays an important role in many life activities such as cell cycle regulation, cell differentiation regulation and epigenetic regulation (Gupta et al., 2010; Wapinski et al., 2011). So far, the research on lncRNA has mainly focused on some model animals and humans, and the research on p...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/113C12N15/82A01H5/00
Inventor 张保龙刘廷利杨郁文陈天子王金彦凌溪铁胡节立阚家亮余文贵
Owner JIANGSU ACAD OF AGRI SCI