Application of Lox (lipoxygenase) D gene of tomato in regulating and controlling plant resistance
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A transgenic plant, plant technology, applied in applications, plant products, genetic engineering, etc.
Active Publication Date: 2014-03-26
INST OF GENETICS & DEVELOPMENTAL BIOLOGY CHINESE ACAD OF SCI
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As early as 1997, the LoxD gene was cloned, but the gene function of LoxD still needs to be studied in more detail
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Embodiment 1
[0029] Embodiment 1, obtain the recombinant vector that suppresses LoxD gene expression
[0030] The total RNA of tomato (Solanum lycopersicon) variety CM was extracted, the cDNA obtained by reverse transcription was used as a template, and PCR amplification was performed with primer 1 and primer 2.
[0031] Primer 1: 5′-GCG CTCGAG AACCAGCAGTTTCAAGGAAGGAGG-3′ (upstream primer, the base underlined is the XhoI recognition site)
[0032] Primer 2: 5′-GCG ACTAGT AACCGGACCACATGCAAACCCCT-3′ (downstream primer, the base underlined is the SpeI recognition site)
[0033] A 465bp PCR product was obtained. After sequencing, the nucleotide sequence of the PCR product was the 106-570th nucleotide from the 5' end of Sequence 2 in the Sequence Listing, which was a partial fragment of the LoxD gene.
[0034] After the above PCR product was double-digested with XhoI and SpeI, it was mixed with the PUCC RNAi vector (recorded in Gan D, Zhang J, Jiang H, Jiang T, Zhu S, Cheng B. Bacterially ...
Embodiment 2
[0037] Example 2, the acquisition and functional identification of LoxD RNAi transgenic tomato
[0038] 1. LoxD RNAi transgenic tomato
[0039] 1. Obtaining recombinant bacteria that inhibit the expression of the LoxD gene
[0040] The recombinant vector pCAMBIA 1301-LoxD RNAi obtained in Example 1 to suppress the expression of the LoxD gene was transformed into Agrobacterium AGLO (recorded by Li C, Liu G, Xu C, Lee GI, Bauer P, Ling HQ, Ganal MW, Howe GA.The tomato suppressor of prosystemin-mediated responses2 gene encodes a fatty acid desaturase required for the biosynthesis of jasmonic acid and the production of a systemic wound signal for defense gene expression. Plant Cell. obtained from the Institute of Genetics and Developmental Biology), picked a single clone, and cultured overnight at 28°C with shaking in LB liquid medium containing kanazyme and rifampicin antibiotics to obtain transformants.
[0041] The transformants were identified by bacterial liquid PCR, using ...
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Abstract
The invention discloses an application of Lox (lipoxygenase) D gene of a tomato in regulating and controlling plant resistance. The nucleotide sequence of a DNA (deoxyribonucleic acid) segment provided by the invention is 106th-570th nucleotides from the tail end of 5' of sequence 2 in a sequence table. The invention also provides an application of LoxD protein and a coding gene thereof in regulating insect resistance and/or disease resistance of plants, wherein the amino acid sequence of LoxD protein is the sequence 1 in the sequence table, the nucleotide sequence of the LoxD protein coding gene is the sequence 2 in the sequence table. Experiments of the invention prove that a transgenic tomato can be obtained by inhibiting expression of a LoxD gene of a wild tomato, the disease resistance and insect resistance of the transgenic tomato are worse than those of the wild tomato, so that inhibition of expression of LoxD genes of plants lowers the insect resistance and disease resistance of the plants, and the LoxD gene is related to disease resistance of plants.
Description
technical field [0001] The invention relates to the field of biotechnology, in particular to the application of a tomato LoxD gene in regulating plant resistance. Background technique [0002] LOX (lipoxygenase, lipoxygenase) is ubiquitous in plants, animals and fungi, and in recent years it has been found that this type of enzyme also exists in bacteria. According to the position of LOX catalyzing the oxidation of diene bonds, it can be divided into 9-lipoxygenase and 13-lipoxygenase; according to whether the LOX protein is located in the chloroplast, LOX can be divided into two categories: Type1 and Type2. In plants, LOX not only participates in their growth and development, but also plays an important role in plant resistance responses such as insect chewing and mechanical damage. There are multiple copies of Lox genes in the plant genomes of tomato, Arabidopsis and other species, and different Lox genes have different biological functions. During JA biosynthesis, Lox s...
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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/11C12N9/02C12N15/53A01H5/00
Inventor 李传友闫留华王保翟庆哲
Owner INST OF GENETICS & DEVELOPMENTAL BIOLOGY CHINESE ACAD OF SCI
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