High-molecular wheat glutelin subunit gene, nucleic acid sequence of promoter thereof, and application thereof

A glutenin subunit, high molecular weight technology, applied in the fields of application, genetic engineering, plant genetic improvement, etc.

Inactive Publication Date: 2007-05-30
SICHUAN AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Since the 1Ay gene is not expressed in common wheat, the effect of this gene on the processing quality of wheat flour has not been reported

Method used

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  • High-molecular wheat glutelin subunit gene, nucleic acid sequence of promoter thereof, and application thereof
  • High-molecular wheat glutelin subunit gene, nucleic acid sequence of promoter thereof, and application thereof
  • High-molecular wheat glutelin subunit gene, nucleic acid sequence of promoter thereof, and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0096] Example 1. SDS-PAGE Analysis of Uraltu Wheat High Molecular Weight Glutenin Subunits

[0097] (1) Plant material: Urartu wheat, common wheat variety China Spring

[0098] (2) Gluten sample preparation

[0099] Take half a seed, weigh it, crush it with sample forceps, put it into a 1.5ml centrifuge tube, then add the sample extract at a ratio of 1mg plus 25μl, boil it in boiling water for 2.5min at room temperature for 2hr, centrifuge at 8000r / min for 5min, Take 7-10 μl of the supernatant, and sample it for analysis. Protein extraction buffer includes: 62.5mM Tris-HCl pH6.8, 2% (W / V) SDS, 10% (V / V) glycerol, 1.5% (W / V) DTT, 0.002% (W / V) bromophenol blue.

[0100] (3) Electrophoresis analysis

[0101] Take Chinese spring (subunit combination: nul1, 7+8, 2+12) as a control, electrophoresis at a constant current of 15-20 mA, after the indicator runs out of the separation gel for 30 minutes, remove the gel and stain overnight, and decolorize the gel with the decolorizati...

Embodiment 2

[0102] Example 2. Isolation and Identification of Uraltu Wheat High Molecular Weight Glutenin Subunit Gene

[0103] (1) Primer design

[0104] According to the published sequence information of the coding region and promoter region of the high-molecular-weight glutenin subunit gene, primers were designed to amplify the coding region of the high-molecular-weight glutenin subunit gene and specifically amplify the promoter region of the y-type subunit gene ,details as follows:

[0105] Primers for amplifying the coding region:

[0106] P 1 : 5′-AGCTGCAGAGAGTTCTATCA-3′

[0107] P 2 : 5′-ATCACCCACAACACCGAGCA-3′

[0108] Primers for amplifying the promoter region:

[0109] P 3 : 5′-AGGGAAAGACAATGGACATG-3′

[0110] P 4 : 5′-CATCTGGAGCCCCGTGCTC-3′

[0111] (2) Genomic DNA extraction by CTAB method

[0112] a) Take 2g of fresh young leaves, grind them into fine powder with liquid nitrogen, add CTAB extract (2% CTAB; 1.4M NaCl; 0.1M Tris-HCl, pH=8.0; 0.1M EDTA, pH =8.0; add ...

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PUM

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Abstract

The invention discloses a coded gene sequence and amino acid sequence with high-expressive activity macromolecular glutelin subunit with sequence as SEQ ID No.1 and SEQ ID No.2 in the biological gene engineering technical domain, which is characterized by the following: providing astopic endosperm promoter with high-expressive activity macromolecular glutelin subunit with sequence as SEQ ID No.3; constructing gene-transmitting carrier; utilizing gene shooting method, agricillin dielectric method and pollen tube channel method to transmit gene to common wheat; culturing superior wheat.

Description

technical field [0001] The invention belongs to the technical field of biogenetic engineering, and in particular relates to a high-molecular-weight glutenin subunit gene 1Ay with expression activity and a nucleic acid sequence of an endosperm-specific expression promoter and application thereof. Background technique [0002] Wheat is one of the most important food crops, accounting for about 17% of the cultivated area of ​​food crops in the world, and is the basic source of human food and nutrition. Wheat endosperm storage proteins can be divided into two categories: Glutenin and Gliadin. According to the difference of SDS-PAGE electrophoretic mobility, gluten can be divided into high molecular weight glutenin subunits (High molecular weight glutenin subunits, HMW-GS) and low molecular weight glutenin subunits (LHW-GS ). High-molecular-weight glutenin subunits are an important storage protein in wheat seeds, and are the main factors that determine the processing and baking...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/29C07K14/415C12N15/82
Inventor 郑有良江千涛魏育明颜泽洪
Owner SICHUAN AGRI UNIV
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