Corn WRKY transcription factor ZmWRKY58, and coding gene and application thereof

A technology of transcription factors and coding genes, which is applied in the fields of genetic engineering and crop genetics and breeding, and can solve the problems of few research reports on other species

Inactive Publication Date: 2013-07-31
ANHUI AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, most of the research on WRKY family transcription factors comes f...

Method used

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  • Corn WRKY transcription factor ZmWRKY58, and coding gene and application thereof
  • Corn WRKY transcription factor ZmWRKY58, and coding gene and application thereof
  • Corn WRKY transcription factor ZmWRKY58, and coding gene and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035] Example 1 Acquisition of Maize WRKY Family Gene ZmWRKY58

[0036] 1. Stress treatment: the plant material is corn B73 inbred line, selected seeds with uniform size and full grains are sown in flower pots with sandy soil, and distilled water is sprayed timely to keep the soil moist. When the seedlings grew to two leaves and one heart, the normal irrigated plants were used as controls, and the stress treatments of 16% PEG-6000 and 200mmol / L NaCl were carried out respectively, and the treatment time was 12, 24, and 48 hours respectively. The scissors cut the leaves of the same part of the maize seedlings in the test group and the control group respectively, and three samples were taken at each treatment stage. After sampling, the samples were immediately frozen in liquid nitrogen and stored in a -80°C refrigerator.

[0037]2. RNA extraction: The corn material obtained above was ground by adding liquid nitrogen, and then quickly transferred to a 1.5 ml centrifuge tube (pre...

Embodiment 2

[0043] Example 2 Sequence Homology and Homology Analysis of Maize ZmWRKY58

[0044] According to the sequence sequencing results, the sequence comparison was performed in the NCBI database, and it was found that the cloned gene sequence had the closest homology relationship with the WRKY family transcription factors. Comparing the protein sequence of this transcription factor with other members of the WRKY transcription factor family, the structure type of its zinc finger is analyzed as C2H2 type. According to the structural characteristics of the DNA binding domain, the ZmWRKY58 protein belongs to the class II WRKY transcription factor family (such as figure 1 ). In order to further analyze the phylogenetic relationship between ZmWRKY58 and other WRKY transcription factors, the phylogenetic relationship between WRKY proteins and ZmWRKY58 in different plants was analyzed (such as figure 2 ).

Embodiment 3

[0045] Example 3 Subcellular localization of ZmWRKY58

[0046] Specific primers were designed with Primer Premier 5.0 software, and the recognition sequences of restriction endonucleases NcoI and SpeI and corresponding protective bases were added to the 5' and 3' ends of the primers, respectively, and handed over to Sangon Biotech. The target fragment was amplified with the correctly sequenced recombinant plasmid P-ZmWRKY58 as a template. The primer sequences are as follows:

[0047] RH-F 5’- ATATCCATGGTAGATGAGGAAGTGGAG-3’

[0048] RH-R 5'-GGACTAGTCACCTGTGCTGCTGCTGC-3'

[0049] The amplified PCR product was recovered and connected to the pMD18-T vector, transformed into Escherichia coli competent by heat shock, a single colony was picked, the plasmid was extracted, and verified by NcoI and SpeI digestion, and the verified correct plasmid was sent for sequencing.

[0050] The above-mentioned recombinant plasmid and pCAMBIA1302 vector were digested with NcoI and SpeI, the tar...

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Abstract

The invention relates to the technical fields of the molecular biology and the gene engineering, and concretely relates to a ZmWRKY58 gene obtained through the amplification in the total cDNA of a corn inbred line B73 by utilizing a PCR technology according to a transcription factor ZmWRKY58 gene sequence design primer. The gene codes a protein having one of amino acid residue sequences comprising SEQ ID No:1 in a sequence table, and an amino acid residue sequence obtained through substituting and/or deleting and/or adding 1-11 amino acid residues to the amino acid residue sequence represented by SEQ ID No:1 and having an interaction with a W box (TTGACC/T) cis-element to improve the stress resistances of plants. Results of onion cuticle subcell localization experiments show that the ZmWRKY58 protein has the nuclear localization characteristic of the transcription factor. The corn ZmWRKY58 gene is used for rice transformation, and obtained transgenic rice has an improved sensitivity to ABA, and has obviously-enhanced salt and drought tolerances. The corn ZmWRKY58 gene provides an important gene resource for the plant stress resistance gene engineering, and is of great significance to the improvement of the output of crops.

Description

technical field [0001] The invention belongs to the fields of genetic engineering and crop genetic breeding, and specifically relates to a coding gene of corn WRKY transcription factor ZmWRKY58 and its application in cultivating stress-tolerant plants. Background technique [0002] Plants often encounter various unfavorable natural environments during the growth process, such as drought, high salinity, low temperature, high temperature flooding, etc. Among them, drought and high salinity are the most important factors that affect plant growth and production and limit the geographical distribution of plants. of the two factors. Drought can cause plant cells to lose water, resulting in complete loss of cell turgor, or even death; high salinity can disrupt the balance of water potential and ion distribution, resulting in molecular damage to plants and delayed growth. During the long evolutionary process, plants have developed a complex set of mechanisms to adapt to abiotic str...

Claims

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Application Information

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IPC IPC(8): C07K14/415C12N15/29C12N15/63C12N1/15C12N1/19C12N1/21C12N5/10C12N15/11A01H5/00
Inventor 朱苏文蔡荣号程备久江海洋林勇翔曹建刚姜翠萍
Owner ANHUI AGRICULTURAL UNIVERSITY
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