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Plant stomata specific promoter and application thereof

A promoter, plant technology, applied in applications, plant products, climate change adaptation, etc., can solve problems such as lack of stomatal expression specificity

Inactive Publication Date: 2012-05-02
CHINA AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

That is, they also lack the specificity of stomatal expression

Method used

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  • Plant stomata specific promoter and application thereof
  • Plant stomata specific promoter and application thereof
  • Plant stomata specific promoter and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0091] Embodiment 1, the cloning and isolation of sea island cotton stomatal-specific promoter SLSP (GbSLSP)

[0092] 1. Extraction of sea island cotton total DNA

[0093] The material was taken from Gossypium barbadense L. variety "SHZ2-214" (purchased from Shihezi University in Xinjiang), 1-2g of young leaves, and the total DNA was extracted by CTAB method (Molecular Cloning, third edition). Take 1-5 μl DNA sample, measure its concentration and purity with a UV spectrophotometer, and use 0.8% agarose gel electrophoresis to detect the DNA purity and integrity. Store the extracted DNA at -20°C.

[0094] 2. PCR amplification and recovery of amplified fragments

[0095] The following two primers (primer 1 and primer 2) were designed and synthesized to amplify the stomatal-specific promoter of Gossypium japonicus. In order to facilitate subsequent cloning and construction, restriction endonuclease Hind III was added to the 5' ends of the two primers and BamH I recognition sequ...

Embodiment 2

[0104] Embodiment 2, sea island cotton stomatal-specific promoter (GbSLSP) drives the construction of GUS gene or GFP gene plant expression vector

[0105] Two-step ligation method: pBS-GbSLSP is double-digested with restriction enzymes Hind III and BamH I, electrophoresed on 1.0% agarose gel, and two target bands of about 650bp and about 350bp are cut out, and DNA fragment recovery reagents are used The box (product of Beijing Dingguo Company) was recovered and purified. The target band (target band 1) of about 650 bp after purification is connected with the large fragment (about 13.9 kb) of the plant expression vector pBI121 plasmid (formerly Clontech company product) that has been digested with Hind III and BamH I to obtain a recombinant plasmid It was named pBI-Gb650; the purified target band (target band 2) of about 350 bp was connected to the recombinant plasmid pBI-Gb650 that had been digested with Hind III to obtain the recombinant plasmid pBI-Gb650+350 for transformat...

Embodiment 3

[0108] Embodiment 3, the acquisition and identification of transgenic GbSLSP tobacco and Arabidopsis

[0109] 1. Transformation of Agrobacterium and identification of transformants

[0110] Competent cells of Agrobacterium tumefaciens strains LBA4404 and GV3101 (products of Life Technology, USA) were prepared by the CaCl2 method (Molecular Cloning, third edition). The two recombinant expression vectors (pBI-GbSLSP-GUS and pBI-GbSLSP-GFP) obtained in Example 2 were transferred into the prepared LBA4404 competent cells by the freeze-thaw method ("Molecular Cloning" third edition), and at the same time, the pBI -GbSLSP-GUS vector was transformed into GV3101 competent cells. The transformed LBA4404 bacterial strain is inoculated into the YEB solid medium containing streptomycin (Str) 100mg / L and kanamycin (Kan) 50mg / L, the transformed GV3101 bacterial strain is inoculated into the YEB solid medium containing rifampicin (Rif) 25mg / L. L and Kanamycin (Kan) 50mg / L YEB solid medium,...

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Abstract

The invention discloses a plant stomata specific promoter and an application thereof. The promoter provided in the invention comprises the following nucleotide sequences: 1) a sequence 1 in a sequence table; 2) a nucleotide sequence still has the function of a stomata specific promoter after the 5' terminal and / or 3' terminal of the nucleotide sequence of the sequence 1 in the sequence table are deleted; and 3) a nucleotide sequence with the function of a stomata specific promoter when one or more deoxynucleotide residues of the nucleotide sequence in 1) or 2) is substituted, added or deleted. Experiments prove that: the promoter provided in the invention has excellent stomata specificity, and the specificity can be stably transferred to the next generation, which has important significance to molecular biological researches of stomata and guard cells, stomata motion and other theories, and has great application value and a wide market prospect in improving the photosynthetic efficiency of crops by culturing drought and disease resisting crops so as to improve yield, quality and other fields.

Description

technical field [0001] The invention relates to a plant stomatal specific promoter and its application. Background technique [0002] Promoter (Promoter) is an important part of genomic genes, it is like a "switch", at the level of transcription, it basically determines whether the coding gene under its control is expressed, when it is expressed, where it is expressed and the expression intensity. Generally, promoters can be roughly divided into three categories according to their mode of action and function: constitutive promoters, specific promoters and inducible promoters (Wang Guanlin, Fang Hongyun, 2002, Principles and Techniques of Plant Genetic Engineering (Second Edition) , Beijing, Science and Technology Press). But this classification is not absolute. In some cases, one type of promoter often has the characteristics of other types of promoters, for example, inducible and enhanced constitutive promoters (Xiao Xingguo et al., Invention Patent No. 200710177962.4). ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/113C12N15/11C12N15/63C12N1/15C12N1/19C12N1/21C12N5/10A01H5/00
CPCY02A40/10
Inventor 肖兴国韩蕾
Owner CHINA AGRI UNIV
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