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Compositions and methods related to silicon transport

A transporter, plant technology, applied in the direction of botany equipment and methods, biochemical equipment and methods, chemical instruments and methods, etc., can solve environmental damage, expensive and other problems

Inactive Publication Date: 2011-09-07
UNIV LAVAL
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] While this rust can be treated with chemical fungicides, doing so is expensive, potentially damaging to the environment, and may only be partially effective

Method used

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  • Compositions and methods related to silicon transport
  • Compositions and methods related to silicon transport
  • Compositions and methods related to silicon transport

Examples

Experimental program
Comparison scheme
Effect test

example 1

[0150] Preparation of total cDNA extracts from wheat roots

[0151] Wheat plant cultivar HY644 was grown in a hydroponic system set to immerse roots in a nutrient solution for 15 minutes every 30 minutes. Each system contained 12 pots, each containing 2-3 seeds grown on vermiculite. These systems were maintained in a greenhouse (16 hours light at 22°C and 8 hours dark at 18°C, 80% humidity). Germinate the seeds in distilled water, replacing the distilled water with Hoagland nutrient solution at the 2-3 leaf stage. This solution was replaced every other week. Once mature, wheat roots were carefully recovered and immediately frozen in liquid nitrogen. Crush the frozen roots in a clean autoclaved mortar and pestle. Total mRNA was then extracted from these root powders using an RNA extraction kit (QIAgen); the RNA was stored at -80°C until use. Add 5 μl of a 300 ng / μl total mRNA to a mixture containing: 2 μl oligodT18 (5 μM), 1 μl of dNTP (10 mM), and 4.5 μl of RNAse-free wat...

example 2

[0153] Amplification and Cloning of a Silicon Transporter Fragment in Wheat

[0154] 100 ng of wheat cDNA obtained as described in Example 1 was added to a mixture containing: 1 μl of dNTP (10 mM), 2.5 μl of pf TP 10X, 1.5 μl of 25 mM MgCl 2 , 12.75 μl of ddH 2 O. 0.25 μl of HotStart Taq DNA polymerase (Eppendorf), 1 μl of 5 μM primer IF (SEQ ID NO: 7), and 1 μl of 5 μM primer 2R (SEQ ID NO: 8). PCR was performed using the following conditions. Initial denaturation at 94°C for 2 minutes, followed by 40 cycles of denaturation (94°C, 30 seconds), annealing (62°C, 30 seconds), and primer extension (72°C, 1 minute), and a final Extension (72°C, 10 minutes).

[0155] Agarose gel electrophoresis of the PCR product demonstrated a unique band of the expected size. 3 μl of this purified PCR product was added to 5 μl of TP 2X Quick Ligation Buffer, 1 μl of 50 ng / μl pGEM-TT plasmid (Promega), and 1 μl of T4 DNA Ligase (Promega) and incubated overnight at 4°C. 5 μl of the ligation re...

example 3

[0159] 3'RACE (Rapid Amplification of cDNA Ends) in Wheat

[0160] 100 ng of wheat cDNA obtained as described in Example 1 was added to a mixture containing: 1 μl of dNTP (10 mM), 2.5 μl of pf TP 10X, 1.5 μl of 25 mM MgCl 2 , 12.75 μl of ddH 2 O. 0.25 μl of HotStart Taq DNA polymerase (Eppendorf), 1 μl of 5 μM DAPT primer (SEQ ID NO: 16), and 1 μl of 5 μM BIeR primer (SEQ ID NO: 18). PCR was performed using the following conditions. Initial denaturation at 94°C for 2 minutes, followed by 40 cycles of denaturation (94°C, 1 minute), annealing (52°C, 30 seconds), and primer extension (72°C, 1 minute), and a final extension (72°C, 10 minutes). The PCR product was diluted 100-fold and another amplification was performed with the same conditions (but using 1 μl of 5 μM ADA primer (SEQ ID NO: 17) and 1 μl of 5 μM BleRNested (5 μM, 5′-CCTGCGAAGATGGAGGTAA-3′) .

[0161] Analysis of the PCR product on agarose gel electrophoresis demonstrated a unique band of this expected size. Th...

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Abstract

Based on our identification of silicon influx and efflux transporter genes in plants known to take up silicon efficiently, including wheat, horsetail, oat, sorghum, and barley, the present invention features polynucleotides encoding silicon transporters; vectors, cells, and plants including such polynucleotides, and methods for making such plants. The invention also features silicon transporter polypeptides and fragments thereof. Plants expressing heterologous silicon transporters may exhibit both increased silicon uptake and increased resistance to biotic and abiotic stresses. In particular, plants such as soybean expressing silicon transporters may exhibit increased resistance to pathogens such as rust.

Description

Background of the invention [0001] The present invention relates to compositions and methods that can be used to increase silicon uptake and increase tolerance to biotic and abiotic stresses in plants such as soybeans. [0002] Biological and abiotic stresses to plants cause billions of dollars worth of damage to crops every year. For example, soybean rust, a disease caused by the fungus Phakopsora pachyrhizi, caused damage worth approximately one billion dollars in Brazil in 2003. The disease has now begun to spread into the United States (the world's largest producer of soybeans). [0003] While this rust can be treated using chemical fungicides, doing so is expensive, potentially damaging to the environment, and may be only partially effective. Therefore, there is a need for additional or improved methods for protecting plants from biotic as well as abiotic stresses. The prevention or control of soybean rust is one of the most important applications in this regard. Sum...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/29A01H5/00A01H5/10C07K14/415C12N15/09C12N15/82C12N5/10
CPCC12N15/8271C07K14/415C12N15/8279C12N15/8282
Inventor 理查德·贝朗葛维尔弗雷德·雷穆斯-博雷尔卡洛琳·格莱戈瓦
Owner UNIV LAVAL
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