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Gene promoter originated from cotton and its application

A promoter and cotton technology, applied in the fields of application, genetic engineering, plant gene improvement, etc., can solve the problems of few structural features of the promoter, no identification of the promoter region of plant glucuronosyltransferase, etc.

Inactive Publication Date: 2009-12-09
TSINGHUA UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although these reports show that glucuronosyltransferases present different expression profiles and functions in plants, the structural characteristics of the promoters that regulate their expression are still poorly understood. Research Report on Identification of Promoter Region

Method used

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  • Gene promoter originated from cotton and its application
  • Gene promoter originated from cotton and its application
  • Gene promoter originated from cotton and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0024] Embodiment 1, cloning and sequence analysis of the promoter pGhGlcat1 of the cotton glucuronosyltransferase gene GhGlcat1

[0025] 1. Cloning of the promoter pGhGlcat1 of the cotton glucuronosyltransferase gene GhGlcat1

[0026] (Paterson AH, Brubaker CL, Wendel JF. A rapid method for extraction cotton (Gossypium spp.) Genomic DNA suitable for RFLP or PCR analysis. Plant Mol Biol Rep 1993; 11:122-7.) was extracted from cotton varieties with reference to the method of Paterson et al. Nuclear genomic DNA from cotton 12 (Gossypium hirsutum cv. CRI 12) leaves was then analyzed with an improved chromosome walking method (A.M.Wu, J.Y.Liu, An improved method of genomic walking for promoter sequences cloning, Chin.J.Biochem.Mol.Biol. 22 (2006) 243-246.) Cloning of the promoter sequence of cotton glucuronosyltransferase gene GhGlcat1. The cloned and purified DNA fragments were ligated into the vector pGEM-T Easy (Promega Company), transformed into E.coli DH5α competent cells, a...

Embodiment 2

[0033] Example 2. Transformation of tobacco with plant expression vector containing pGhGlcat1 promoter and GUS gene and determination of GUS activity and histochemical localization

[0034] 1. Construction of plant expression vector pBI-pGhGlcat1::GUS containing pGhGlcat1 promoter and GUS gene

[0035] In order to detect the activity of the GhGlcat1 promoter pGhGlcat1 in transgenic tobacco, a tobacco expression vector containing pGhGlcat1 and GUS genes is now constructed. The specific method is: using the recombinant vector pGEM-T Easy-pGhGlcat1 constructed in Example 1 as a template, in Forward primer 1: 5'-C AAGCTT CAGACCTGAGTCATTC-3' (underlined base is restriction endonuclease Hind III recognition site) and reverse primer 2: 5'-CT GGATCC Under the guidance of CTTATGAGTAAAATGGAATT-3' (the underlined base is the recognition site of restriction endonuclease BamHI), pGhGlcat1 was amplified by PCR, and the recognition sites of restriction endonuclease Hind III and BamHI were...

Embodiment 3

[0048] Example 3, Detection of the impact of pGhGlcat1 transgenic tobacco on abiotic stress

[0049] The T2 generation pBI-pGhGlcat1::GUS transgenic tobacco grown in the greenhouse for 3 weeks was treated with abiotic stress, and the roots of the plants were respectively immersed in the following solutions using MS culture medium as solvent: 200mM sucrose (Suc), 200mM glucose (Glu), 200mM fructose (Fru), 200mM mannitol (Man), 200mM sorbitol (Sor), 50μM naphthaleneacetic acid (NAA), 10μM gibberellin (GA), 50μM abscisic acid (ABA), 20% PEG8000 ( PEG), 250mM NaCl, 50μM methyljasmonic acid (MeJA) and 1mM ethylene (Eth), placed in an incubator at 25°C for 24h, and then the GUS activity was measured. Set MS culture medium as the control (CK), all GUS activity Data are the mean ± SE (standard error) of values ​​determined for 5 different transgenic lines.

[0050] The GUS relative activity analysis results of the transgenic plants treated with the above-mentioned different solution ...

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Abstract

The invention discloses a plant gene promoter and its application. The base sequence of the promoter is shown in SEQ ID NO:1. Tobacco transgenic experiments proved that pGhGlcat1 can endow high-level expression of GUS reporter gene in epidermal trichomes, apical meristem region, and stamens and pistils of plant floral organs, indicating that GhGlcat1 plays a role in cotton fiber primary cell wall synthesis, cell elongation, floral organ development and stress The functional study of pGhGlcat1 provides clues for revealing the expression regulation mechanism of GhGlcat1. In addition, pGhGlcat1 can also be used in the genetic engineering improvement of plant quality and resistance.

Description

technical field [0001] The present invention relates to a plant gene promoter and its application, in particular to a plant gene promoter derived from cotton and its application in plant quality improvement. Background technique [0002] Glycosyltransferases (GTs) catalyze the attachment of sugar groups to different acceptor molecules. Based on their sequence similarity, catalytic specificity, conserved sequence and donor sugar species, these enzymes are divided into different families ( T.Vogt, P.Jones, Glycosyltransferases in plant natural products synthesis: characterization of a supergene family, Trends Plant Sci.5(2000) 380-386; W.G.T.Willats, L.McCartney, W.Mackie, J.P.Knox, Pectin: cellbiology and prospects for functional analysis. Plant Mol. Biol. 47(2001) 9-27.). In plants, glycosyltransferases can convert the products of photosynthesis into disaccharides, oligosaccharides, polysaccharides and other small sugar molecules, so they play an important role in the devel...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/29C12N15/63C12N5/10C12N15/82
Inventor 刘进元吴蔼民吕世友
Owner TSINGHUA UNIV
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