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Peanut C2H2 type salt-resistance zine finger protein gene AhZFP1 and coded protein and gene cloning method thereof

A technology of C2H2 and zinc finger protein, which is applied in the field of crop salt tolerance gene research to achieve the effect of increasing expression

Inactive Publication Date: 2010-03-17
SHANDONG PEANUT RES INST
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] At present, there have been many papers on Arabidopsis, rice, corn, wheat and other crops. 2 h 2 It is reported that the type zinc finger protein gene improves the salt tolerance of plants, but there is no related report in peanut. Therefore, for peanut C 2 h 2 The cloning of type zinc finger protein genes and the study of their biological functions will help to clarify the molecular mechanism of peanut salt tolerance, and at the same time obtain genetic resources that can be used to improve peanut salt tolerance

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0023] Example 1: Peanut C 2 h 2 Molecular cloning of type zinc finger protein gene AhZFP1

[0024] Luhua 28 peanut seeds were selected as the experimental material. After being induced by salt, the total RNA of peanut tissues was extracted with Trizol reagent (Tiangen) at room temperature, and treated with DNase I (Takara) to remove genomic DNA contamination in the samples. cDNA was obtained by reverse transcription using Promega's RT-PCR system. The primers for coding region of peanut AhZFP1 gene were designed according to the EST information in NCBI database: forward primer: 5'-CAC TAC TCT GTT CGT ACT CTG-3', reverse primer: 5'-TGC TCT TCA ACCTTC TTC-3'. Takara's Ex-Taq enzyme was used for PCR amplification, and the PCR product was recovered and ligated with MD18-T simple vector (Takara). The ligated product was transformed into E.coli Top 10 competent cells, and positive clones were screened by the blue-white method. The screened positive clones were further verified by...

Embodiment 2

[0025] Example 2: Sequence information and characteristic analysis of AhZFP1

[0026] The AhZFP1 gene of the present invention is 835bp long, and its open reading frame is 693bp, located at 61-753bp. BioXM software analysis showed that AhZFP1 encoded a total of 231 amino acids. The bioinformatics analysis of the sequence showed that the protein had a molecular weight of 24.75KDa and contained two C 2 h 2 type zinc finger protein domain. Sequence comparison analysis also showed that AhZFP1 and soybean, rice, petunia, Arabidopsis and other C 2 h 2 There is a high similarity between the two types of zinc finger proteins, especially the conserved regions.

Embodiment 3

[0027] Example 3: Expression study of AhZFP1

[0028]The partial sequence of peanut AhZFP1 gene was used to design primers, and semi-quantitative RT-PCR technology was used to analyze the changes of the gene expression and the expression differences in different peanut tissues under salt stress. The results showed that under the condition of salt induction, the expression level of this gene in roots, stems and leaves all increased, but the induction speed in stems and leaves was higher than that in roots. The analysis results showed that the expression of AhZFP1 gene responded to salt stress, and it was inferred that it played a certain role in the process of salt resistance.

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Abstract

The invention discloses a C2H2 type zine finger protein gene AhZFP1 gene order in peanut, responding to salt stress, and coded protein order thereof. The invention also provides a method for cloning the C2H2 type zine finger protein gene AhZFP1 gene. The expression analysis of the C2H2 type zine finger protein gene AhZFP1 shows that the expression of the gene is obviously increased in different tissues of the peanut under the condition of salt stress, and a transgene test analysis also shows that the overexpression of the gene can surely enhance the salt resistance of the peanut.

Description

technical field [0001] The invention relates to the field of genetic research on crop salt tolerance, in particular to a peanut C 2 h 2 Type salt-resistant zinc finger protein gene AhZFP1 and its encoded protein and gene cloning method. Background technique [0002] Peanut is an important oil crop and economic crop in my country. From the perspective of the production and supply of domestic edible vegetable oil, peanut oil accounts for 25% of the total. It is the only bulk oil that does not need to be imported in my country, and it is the main oil crop that guarantees the supply of oil in my country. In recent years, with the increasing shortage of oil resources and energy resources in the world, more and more attention has been paid to the production of peanuts. Expanding the planting area of ​​peanuts has become an important way to solve the shortage of oil resources in my country. There are 36 million hm2 of saline-alkali land in my country, and more than 2 million hm ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/29C12N15/10C07K14/415
Inventor 禹山林江燕杨庆利陈明娜潘丽娟朱凤任增凯
Owner SHANDONG PEANUT RES INST
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