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Barley Transcription Factor hvbzip10 Gene and Its Application in Wheat Resistance to Stripe Rust and Leaf Rust

A technology for wheat stripe rust and wheat leaves, applied in the fields of application, genetic engineering, plant genetic improvement, etc., can solve the problems of easy loss of resistance and high mutation frequency, and achieve the effect of improving disease resistance

Active Publication Date: 2022-06-24
HEBEI AGRICULTURAL UNIV.
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Both wheat stripe rust and leaf rust pathogens have the characteristics of high mutation frequency, and their physiological races can complete multiple mutations in a short period of time, which makes it easy for single-resistant wheat varieties to lose resistance in a short period of time

Method used

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  • Barley Transcription Factor hvbzip10 Gene and Its Application in Wheat Resistance to Stripe Rust and Leaf Rust
  • Barley Transcription Factor hvbzip10 Gene and Its Application in Wheat Resistance to Stripe Rust and Leaf Rust

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0021] Example 1. Cloning of barley HvbZIP10 gene

[0022] Barley Leaf RNA Extraction: RNA extraction was performed using the RNA Extraction Kit (QIAGEN, Hilden, Germany). The barley material "Golden Promise" seedling stage second leaf leaf sample was quickly ground into powder with liquid nitrogen in a sterilized mortar and ready to use. To prepare the Buffer RLT mixture, add 10 μL of β-mercaptoethanol per ml of Buffer RLT, prepare and use now, and place on ice after mixing. Take out the ground RNA sample from liquid nitrogen, quickly add 500 μL of Buffer RLT mixture, shake well, and centrifuge at 10,000 g for 2 min. Transfer the supernatant to a purple spin column with a pipette and centrifuge at 10,000 g for 1 min. Transfer the collected liquid to the pink spin column, add pre-cooled absolute ethanol (the amount added is 1 / 2 of the collected liquid), invert and mix, and let stand to precipitate the nucleic acid. Immediately leave for 30 s, and then discard the collection...

Embodiment 2

[0030] Example 2. Construction of HvbZIP10 gene wheat transgenic vector pLGY-02

[0031] Construction of wheat transgenic vector pLGY-02: Extract the correctly sequenced HvbZIP10-T recombinant plasmid and the plasmid of pLGY-02 vector, and perform double digestion with restriction enzymes KpnI+SpeI. The specific digestion system is: plasmid 1.0μg, KpnI (15U / μL) 1.0μL, SpeI (10U / μL) 1.0μL, 10×Buffer 2.0μL, ddH 2 O supplemented to 20 μL. The digestion mixture was digested in a metal bath at 37°C for 3-5h. After the digestion products were detected by electrophoresis, the target gene fragment and the pLGY-02 vector fragment were recovered by gel and ligated. Ligation system: 12 μL of target fragment, 5 μL of pLGY-02 vector fragment, 1.0 μL of T4 DNA ligase, 2.0 μL of T4 DNALigase buffer, ddH 2 O supplemented to 20 μL. Centrifuge to mix the reagents well, connect at 22°C for at least 1 h or overnight at 4°C. The ligation product was transformed into E. coli, tested by PCR, pi...

Embodiment 3

[0032] Example 3. Preparation of HvbZIP10 overexpressing wheat transgenic plants

[0033] The preparation of Agrobacterium-mediated wheat transgenic materials was completed by Shandong Jinan Bangdi Biological Co., Ltd. The transformation background material was common wheat spring wheat material JW1, and the method of Agrobacterium-mediated wheat immature embryo transformation was adopted.

[0034] Genomic DNA extraction by SDS method: sample and label; add 1 grinding bead to each tube, pre-cool with liquid nitrogen, balance and put it in the proofing machine, grind at 1100g for 1min, take it out and put it in 600μL Extraction buffer (100mL 0.1M Tris-HCl pH= 7.5, 100mL of 0.5M EDTA pH=8.0, 125mL of 10% SDS), placed in a water bath at 65°C for 30min after shaking; taken out and placed on ice for 15min to cool to room temperature, then added 300μL of 6M Ammonium Acetate (ammonium acetate), mixed well and put in 4 ℃ refrigerator for 15 minutes, 12000g centrifugation for 15 minute...

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Abstract

The invention provides a barley transcription factor HvbZIP10 gene and its application in wheat resistance to stripe rust and leaf rust. The invention relates to gene sequences, and specifically discloses barley transcription factor HvbZIP10, the nucleotide sequence of which is shown in SEQ ID No.1, and the preparation process of wheat transgenic materials overexpressing HvbZIP10. The invention verifies through experiments that the HvbZIP10 gene can significantly improve the resistance level of wheat to wheat stripe rust and wheat leaf rust.

Description

technical field [0001] The invention belongs to the technical field of biological genetic engineering, and relates to a barley transcription factor HvbZIP10 gene and its application in resistance to stripe rust and leaf rust of wheat. Background technique [0002] As a major food crop, the quality and yield of wheat seriously affect my country's food security and social stability. The high and stable yield of wheat is of great significance to my country's agricultural development. Wheat stripe rust and wheat leaf rust caused by Puccinia striiformis f.sp.tritici and Puccinia triticina (Pt), respectively, are important fungal diseases that seriously affect wheat production in my country. . In recent years, due to the increase of planting density and the change of agricultural farming system, the occurrence of wheat stripe rust and leaf rust has become more and more serious, which seriously affects the grain yield and quality in my country. Both the wheat stripe rust and leaf ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/29C12N15/10C12N15/82A01H5/00A01H6/46
CPCC07K14/415C12N15/8282
Inventor 王逍冬苏君陈欣池任小鹏何佳怡尚小凤于秀梅
Owner HEBEI AGRICULTURAL UNIV.
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