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ADP-glucose pyrophosphorylase mutant, encoding genes thereof and application of same two

A technology of encoding genes and genes, which is applied in the fields of application, genetic engineering, plant genetic improvement, etc., and can solve problems such as the discovery of disulfide bond structures

Inactive Publication Date: 2009-11-18
CHINA AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, no disulfide bond structure formed by cysteine ​​was found on the large and small subunits of AGPase in maize endosperm

Method used

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  • ADP-glucose pyrophosphorylase mutant, encoding genes thereof and application of same two
  • ADP-glucose pyrophosphorylase mutant, encoding genes thereof and application of same two
  • ADP-glucose pyrophosphorylase mutant, encoding genes thereof and application of same two

Examples

Experimental program
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Embodiment 1

[0042] Example 1. Obtaining of Maize Endosperm ADP-Glucose Pyrophosphorylase Mutant and Its Encoding Gene

[0043] 1. Obtaining the large and small subunit cDNA of maize endosperm AGPase

[0044] 1. Primer design

[0045] Two pairs of RT-PCR primers (Sh and Bt) were designed according to the cDNA sequences of the large subunit gene Shrunken-2 and the small subunit gene Brittle-2 of maize endosperm AGPase (GenBank numbers: M81603, AF334959, respectively). At the same time, in order to facilitate the construction of the vector, a special design was made for the following two points when designing the primers: a. A recognition site for the restriction endonuclease Nde I was added to the 5' end of Shrunken-2, and a restriction endonuclease was added to the 3' end The recognition sites of enzymes Sma I and SacI; b. Because there is a restriction endonuclease Nco I recognition site at the 5' end ATG position of Brittle-2, then Brittle-2 is added from the 39th A at the 5' end Mutat...

Embodiment 2

[0074] Example 2, Determination of Glycogen Content of Corn Endosperm AGPase Mutant in Transgenic Escherichia coli

[0075] 1. Construction of maize endosperm AGPase mutant vector

[0076] The small subunit gene Bt2 of maize endosperm ADP-glucose pyrophosphorylase mutant genes AGPase-120-33 and AGPase-10-3-53 was artificially synthesized respectively, and NcoI restriction site was introduced into its 5' end respectively. A Sac I restriction site was introduced at the 3' end. The small subunit gene Bt2 of the synthetic maize endosperm ADP-glucose pyrophosphorylase mutant gene AGPase-120-33 and AGPase-10-3-53 was double-digested with Nco I and Sac I, respectively, and the digested products were Agarose gel electrophoresis and recovery of the target band. The vector pGEX-KG-Sh2-Bt2 constructed in Example 1 was double digested with Nco I and Sac I at the same time, and the 6.5Kb large fragment after removing the small subunit gene Bt2 was gel cut and recovered. Then use T4 DNA ...

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Abstract

The invention discloses a corn endosperm ADP-glucose pyrophosphorylase mutant, encoding genes thereof and application of the same two. The protein can be the protein (1) formed by the first amino acid sequences showed as in a sequence table and can also be the protein (2) which is formed by substituting and / or deleting the first amino acid residue sequences showed as in a sequence table and / or adding one or several amino acid residues to the first amino acid residue sequences, has ADP-glucose pyrophosphorylase activity and is derived from the protein (1). The corn endosperm ADP-glucose pyrophosphorylase mutant and the encoding thereof play important roles in the corn variety improvement.

Description

technical field [0001] The invention relates to a plant ADP-glucose pyrophosphorylase mutant, its coding gene and application, in particular to the ADP-glucose pyrophosphorylase mutant derived from corn endosperm, its coding gene and its application. Background technique [0002] As an irreplaceable material resource in industrial production and human life, starch plays an important role in human production and life. Therefore, researchers have devoted great enthusiasm to the study of starch synthesis and have never stopped. For a long time in the past, in order to meet the food demand and industrial raw material supply brought by the continuously growing world population, people only devoted themselves to increasing the yield of crops (such as corn, wheat, rice, etc.). However, with the rapid development of society and the continuous improvement of people's living standards, and from the perspective of environmental protection and sustainable development, the requirements f...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N9/12C12N15/54C12N15/63C12N5/10C12N1/15C12N1/19C12N1/21C12N15/11C12N15/82A01H1/00A01H5/10
Inventor 王建华王国英林衡
Owner CHINA AGRI UNIV
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