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Maize zmstp1 protein and its coding gene and application

A gene and corn technology, applied in the field of molecular biology, can solve problems such as different monosaccharide absorption capacity, and achieve the effect of improving plant nitrogen efficiency

Active Publication Date: 2020-03-06
CHINA AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, so far, sugar transporters in maize have rarely been reported. In this study, the sugar transporter STP1 gene was cloned from maize and complemented to yeast to express glucose (Glc), fructose (Frc), mannose (Mannose). ) a strong response, and a partial response to galactose (Gal), indicating that ZmSTP1 can absorb a variety of monosaccharides after being transferred into the yeast system, but the absorption capacity of different monosaccharides is different

Method used

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  • Maize zmstp1 protein and its coding gene and application
  • Maize zmstp1 protein and its coding gene and application
  • Maize zmstp1 protein and its coding gene and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0056] Embodiment 1, the cloning of corn sugar transporter gene ZmSTP1 gene

[0057] The amino acid sequences of monosaccharide transporters in Arabidopsis thaliana, rice, and wheat were obtained from NCBI, Maizesequence, and Uniprot databases, and compared using ClustalW1.8 (Thompson et al. 1994 Nucleic Acids Research 22, 4673-4680). Phylogenetic tree analysis showed that the genetic relationship with AtSTP1 was the closest, so the gene was named ZmSTP1 ( figure 1 ).

[0058] 1. Total RNA extraction

[0059] Take 200mg of fresh B73 corn roots and grind them in liquid nitrogen; add 1ml of Trizol extract provided in the kit, shake at room temperature for 5 minutes; then add 200μl of chloroform, shake for 30 seconds, centrifuge at 12,000 rpm for 15 minutes at 4°C; Add 0.5ml of isopropanol, let stand at room temperature for 1 hour, centrifuge at 12,000 rpm at 4°C for 15 minutes; take the precipitate, add 1ml of 70% ethanol, shake for 1 minute, and centrifuge at 10,000 rpm at 4°...

Embodiment 2

[0080] Example 2, Real-Time PCR analysis of the expression characteristics of various tissues of the maize ZmSPT1 gene.

[0081] The test materials were planted at the Shangzhuang Experimental Station of China Agricultural University. Corn samples of different tissues were harvested one week after silking, quickly placed in liquid nitrogen, and brought back to the laboratory and placed in a -80 degree refrigerator for later use.

[0082] Quantitative results showed that ZmSTP1 had the highest expression level in roots ( figure 2 ).

[0083] Attached Real-Time PCR operation steps:

[0084] 1, extract the total RNA in different processing sample roots (method is with embodiment 1);

[0085] 2. Take 50 μg of total RNA, and use DNase I (TaKaRa Company, catalog number: D2215) to remove genomic DNA, as follows:

[0086] Reaction system (50μl):

[0087]

[0088] React at 37°C for 30 minutes;

[0089] Add 150 μl DEPC water, add 200 μl phenol / chloroform / isoamyl alcohol (25:24:...

Embodiment 3

[0103] Example 3. ZmSPT1 expression localization analysis using in situ hybridization and green fluorescent protein technology.

[0104] 1. Preparation of Plant Material

[0105] Hoagland medium was used for plant culture, and root tip samples were taken from corn seedlings at the three-leaf stage. Cut root tip 0.5-1cm and put it into FAA fixative solution (composition per 100ml fixative solution contains: 50% ethanol 90ml, glacial acetic acid 5ml, formaldehyde 5ml);

[0106] The plant material is then dehydrated, cleared and dipped in wax as follows:

[0107] Discard the FAA fixative, wash with DEPC twice;

[0108] 50% ethanol, 50% ethanol+10% tert-butanol, 50% ethanol+20% tert-butanol, 50% ethanol+35% tert-butanol, 50% ethanol+50% tert-butanol, 25% ethanol+75% tert-butanol, 25% ethanol+75% tert-butanol+0.1% eosin (Y), and 100% tert-butanol were treated successively for 2 hours;

[0109] Transfer to 2 / 3 tert-butanol + 1 / 3 paraffin oil and place for 4 hours;

[0110] Pour...

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Abstract

The invention relates to the field of molecular biology, and particularly provides corn ZmSTP1 protein (shown as SEQ ID No.1) and application of coding gene (shown as SEQ ID No.2) of the corn ZmSTP1 in promoting monosaccharide absorption of plant root systems. Protein having a sugar absorbing function is obtained from corn which is an important crop for the first time, and the gene is expressed in root tips of corn and has functions of absorbing and transferring multiple kinds of polysaccharide; by transferring the gene into arabidopsis thaliana which is a model plant, absorbing capability of transgenic plants to specific sugar can be improved, biomass and seed yield of soil-culture plants can be increased remarkably, and the gene has important application prospect; from the perspective of increasing nitrogen efficiency of carbohydrate, the gene is expected to increase plant nitrogen efficiency and cultivate high-nitrogen-efficiency crops.

Description

technical field [0001] The invention relates to the field of molecular biology, in particular to the application of corn ZmSTP1 protein and its coding gene in promoting the monosaccharide absorption of plant roots. Background technique [0002] Carbohydrates, also known as sugar compounds, are the most important and widely distributed organic compounds in nature. Sugar is the product of photosynthesis and the substrate of respiration, which provides carbon skeleton and energy for physiological and biochemical reactions in plants. Therefore, the supply of sugar is very important for carbon and nitrogen metabolism, dry matter accumulation, and crop yield formation. Part of the sugar in plants comes from photosynthesis and starch degradation, and part of it is absorbed directly from the soil by roots. Among them, the sugars directly absorbed by the roots are of great significance to the accumulation of total carbohydrates in plants. Glucose, fructose, sucrose and other neutra...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K14/415C12N15/29C12N15/82A01H5/00A01H6/20
Inventor 李学贤韩洁楠郑红艳
Owner CHINA AGRI UNIV
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