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Stress resistance ERF transcription factor gene derived from Brassica napus

A Brassica napus, transcription factor technology, applied in genetic engineering, plant gene improvement, peptide source, etc., can solve the problems of long time, ineffectiveness, complex conventional breeding of stress resistance genetic basis, etc.

Inactive Publication Date: 2010-06-16
SHANGHAI ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Improving the stress resistance of rapeseed varieties is still one of the important breeding goals in rapeseed genetics and breeding, but it takes a long time to transfer and recombine stress resistance target genes between varieties and species by means of hybridization and backcrossing , coupled with the complexity of the genetic basis of stress resistance makes conventional breeding difficult

Method used

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  • Stress resistance ERF transcription factor gene derived from Brassica napus
  • Stress resistance ERF transcription factor gene derived from Brassica napus

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0052] Example 1: RNA Extraction and cDNA Synthesis of Double Low Cabbage Type Rapeseed Huyou 15 Seedlings

[0053] (1) Test method:

[0054] 1. Extraction of RNA

[0055] Add 100mL extraction buffer (rape RNA extraction buffer formulation: CTAB 3% (W / V); PVP 3% (W / V) (Mw 4000); EDTA 25mM; NaCl 2.0M; Tris-HCl100mM, pH 8.0; Spermidine 0.5g / L; DEPC 0.1% (V / V); 0.1% DEPC-treated SDS 0.5% (W / V); 0.1% DEPC-treated LiCl 10M) into a 50mL polypropylene tube, preheated at 65°C;

[0056] Weigh 5g of plant material and pour it into liquid nitrogen to keep the material in a frozen and brittle state, and grind it;

[0057] After grinding, transfer the fine powder to a 50mL centrifuge tube with preheated extraction buffer at 65°C;

[0058] Place the centrifuge tube in a 65°C water bath for 45 minutes, and shake occasionally to mix the ingredients;

[0059] Add an equal volume of chloroform-isoamyl alcohol mixture, and mix gently up and down for about 10 minutes;

[0060] Centrifuge at ...

Embodiment 2

[0076] Example 2: PCR method to obtain the BnaERFB1-2-Hy15 gene fragment of Brassica napus stress-resistance-related transcription factor

[0077] (1) Test method:

[0078] In the present invention, a pair of primers (BnaERFB1-2-F and BnaERFB1-2-R) are designed, and BamHI and SacI enzyme cutting sites are respectively introduced at both ends of the primers for the needs of construction such as cloning identification.

[0079] PCR reaction system: 5.0 μL of 10×PCR buffer; 4 μL of dNTPs (each 2.5 mM); 1 μL (20 ng) of cDNA template of rape Huyou 15; primer BnaERFB1-2-F 0.5 μL; primer BnaERFB1-2-R 0.5 μL; Ex-Taq 0.4 μL (added after pre-denaturation); add sterile water to make up to 50 μL. PCR reaction program: Denaturation at 94°C for 30 s, annealing at 55°C for 30 s, extension at 72°C for 1 min, a total of 30 cycles of amplification, and extension at 72°C for 10 min.

[0080] (2) Test results:

[0081] Through 1.0% agarose gel electrophoresis, the amplified product is detected...

Embodiment 3

[0082] Embodiment 3: clone identification, sequence determination

[0083] (1) Test method:

[0084] The amplified fragment was recovered by DNA agarose gel recovery kit of Hangzhou Weitejie Biochemical Technology Co., Ltd., and then cloned into the pMD-18-Simple T vector of Dalian Bao Bioengineering Co., Ltd. for cloning identification and sequence determination.

[0085] The present invention finally obtains the BnaERFB1-2-Hy15 gene of the stress-resistance-related transcription factor BnaERFB1-2-Hy15 in Brassica napus through nucleotide sequence determination and analysis, which has the following base and amino acid sequence information.

[0086] (2) Test results:

[0087] Sequencing analysis results showed that the BnaERFB1-2-Hy15 gene coding reading frame of Brassica napus stress-resistance-related transcription factor consisted of 678bp, encoding a protein of 225 amino acids.

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Abstract

The invention provides a stress resistance ERF transcription factor gene derived from Brassica napus, a preparation method and usage thereof. The Brassica napus stress resistance ERF transcription factor gene is BnaERFB1-2-Hy15, the base sequence thereof is shown as SEQ ID No1. In the invention, the gene sequence of Brassica napus stress resistance ERF transcription factor is cloned from Brassica napus seedling by polymerase amplification technology, the obtained ERF transcription factor gene can be used in plant transformation to improve stress resistance of plants.

Description

technical field [0001] The invention relates to the field of crop genetics and breeding, in particular to a stress-resistance-related gene sequence derived from Brassica napus. Background technique [0002] Plants are exposed to a variety of abiotic stresses throughout their life, such as drought, high salinity, and extreme temperature. These stresses have adverse effects on plant growth and development, and ultimately affect their yield and quality. Plants sense stress signals, through a series of signal transductions, and finally start the expression of related genes to help plants adapt to or resist adversity. The products of related gene expression play a role in stress tolerance and response, some of which are only induced by water stress, and some Genes are induced only by low temperature, some genes are induced by both water and low temperature stress (Shinozaki and Yamaguchi, Curr Opin Plant Biol, 2000, 3: 217-22; Yamaguchi and Shinozaki, Annu Rev Plant Biol, 2006, 5...

Claims

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Application Information

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IPC IPC(8): C12N15/29C12N15/10C07K14/415A01H1/00
Inventor 熊爱生姚泉洪庄静彭日荷薛永高峰付晓燕李贤田永生赵伟朱波金晓芬
Owner SHANGHAI ACAD OF AGRI SCI
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