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Method for promoting plant growth and increasing cellulose content and/or yield

A cellulosic, plant-based technology used in bioengineering to increase growth rate and yield

Inactive Publication Date: 2011-08-17
HENAN ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Goldschmidt and Huber (Goldchmidt, E.E. and S.C. Huber.1992.Regulation of Photosynthesis by End-Product Accumulation in Leaves of Plants Stroing Starch, Sucrose and Hexose Sugars.Plant Physiol.99: 1443-1448) carried out on the influence of girdling crop leaves tested and showed that increases in starch and other photosynthetic products did indeed suppress photosynthetic yield

Method used

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  • Method for promoting plant growth and increasing cellulose content and/or yield
  • Method for promoting plant growth and increasing cellulose content and/or yield
  • Method for promoting plant growth and increasing cellulose content and/or yield

Examples

Experimental program
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Effect test

Embodiment 1

[0051] Preparation of recombinant DNA

[0052] The original bacteria (Acetobacter xylinum) contains the UDPG pyrophosphorylase gene (UDPG-PPase), obtained from ATCC (23768). The UDPG pyrophosphorylase gene was cloned after PCR amplification. The design of PCR primers takes into account the following factors:

[0053] Add restriction enzyme sites suitable for cloning into various vectors.

[0054] The start codon was mutated from valine to methionine.

[0055] The Eco RI site is mutated, and the amino acid sequence is not changed after removal.

[0056] A non-coding DNA fragment is added to the 5' end of the gene to increase translation efficiency.

[0057] As a result, the primer A at the 5' end was:

[0058] (MVKPLKKAVL) (SEQ ID NO: 2)

[0059] ta GGATCC gtcgaccATGGTCAAGccccttaaaaaagccgtattgc (SEQ ID NO: 1)

[0060] The 5' end of the original UDPG pyrophosphorylase gene is:

[0061] ttgaggtaaatattaGTGATTAAgccccttaaaaaagccgtattgccggttg → (SEQ ID NO: 3)

[0062] VIKP...

Embodiment 2

[0085] Characteristics of Tobacco Transformation and Expression of UDPG Pyrophosphorylase

[0086] convert

[0087] The binary vector pBLAX6 containing the AxUDPG pyrophosphorylase gene was transformed into Agrobacterium strain EHA 105 and used to infect leaf discs of tobacco c.v.xanthii. More than 42 independent T. 0 Transformed plants are regenerated and individual plants are transferred from the tissue culture chamber to the growth chamber for seed production. UDPG pyrophosphorylase gene in T 0 Stable transformation of tobacco plants has been demonstrated for the first time by PCR amplification using intrinsic primers (see previous report). Further Southern blot analysis was performed. More than 42 independently transformed plants (T 0 ) take root and grow in sterile MS medium and soil. Harvest to get 24 T 0 The seeds of the plant are then used to produce T 1 plants. T 1 Plants were planted in soil after germination on a medium containing kanamycin (150 mg / ml). k...

Embodiment 3

[0098] Protein Expression Analysis

[0099] Protein Production and Antigen Purification

[0100] The protein expression vector pMALAX was used for the overproduction of UDPG pyrophosphorylase-maltose binding protein (MBP), which was integrated into E. coli after induction with isopropylthiogalactopyranoside (IPTG). Crude protein extracts were obtained in a buffer of guanidine and urea. The UDPG pyrophosphatase-MBP fusion protein was purified by affinity chromatography using an amylose resin affinity chromatography column, and the purified protein was eluted from the column with 10 mM maltose. The purified fusion protein was proved to be a pure fragment using SDS-PAGE and rabbit antibodies.

[0101] antibody production

[0102] After immunization of rabbits with purified UDPG pyrophosphorylase-MBP fusion protein, anti-UDPG pyrophosphorylase antibody was produced by Enz-Probe Biotechnology (Enz-Probe Biotechnology, Burnaby, B.C.). UDPG pyrophosphatase-specific antibodies wer...

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Abstract

The invention provides a method for promoting the plant growth and increasing the cellulose content and / or yield and belongs to the field of bioengineering. The method comprises the step of introducing a DNA sequence capable of changing a level of a cellulose precursor into a plant. The cellulose precursor is UPDG (uridine diphosphoglucose). The DNA sequence encodes an enzyme responsible for synthesis of uridine diphosphoglucose, preferably encodes a sequence of UPDG pyrophosphorylase; and is derived from any organisms including plants, bacteria or yeasts, preferably Bacillus xylinus. The expression of the uridine diphosphoglucose pyrophosphorylase gene can change the level of the cellulose precursor in plant tissues or cells, so as to promote the growth of plants and increase the cellulose content and / or yield.

Description

technical field [0001] The invention belongs to the field of bioengineering, and in particular relates to a method for accelerating plant growth and increasing cellulose amount and / or output. Background technique [0002] Cellulose is one of the main functions and components of the cell wall. Taylor (Taylor NG, Howells RM, Huttly AK, Vickers K, Turner SR.2003. Interactions among three distinct CesA proteins essential for cellulose synthesis. Proc Natl Acad Sci USA 100, 1450-1455) and Liang Haiyong et al. Yong, Xia Xiuying, Gao Xiaorong, Su Qiao. 2007. Transformation and expression analysis of antisense 4CL and UGPase bivalent gene in tobacco. The research results of Botany Bulletin 24(4): 459-464) all showed that the cellulose content and positively correlated with cell wall thickness. Dou Yongxiu (Dou Yongxiu. 2008. Evaluation of rice lodging resistance at the fruiting stage and research on the impact of lodging on yield and quality. Yangzhou: Yangzhou University.) Resear...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/09C12N15/82
Inventor 薛保国杨丽荣雷振生全鑫孙虎吴政卿曹岳恩赵献林武超刘明源
Owner HENAN ACAD OF AGRI SCI
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