Peanut MYB transcription factor AhMYB31 and application thereof

A transcription factor, R2R3-MYB technology, applied in application, genetic engineering, plant gene improvement, etc., can solve problems such as functional analysis that has not been reported

Inactive Publication Date: 2016-01-06
JIANGSU ACADEMY OF AGRICULTURAL SCIENCES
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Such transcription factors in peanuts are rarely reported. According to the conservation of the MYB domain, Shandong Peanut Institute Chen et al. (2013) used biological software to compare and analyze the 36,741 ESTs sequences published in peanuts, and obtained 30 sequences with complete ORFs. The cDNA sequence of (open reading frame) was named AhMYB1~30. According to the structural domain analysis, 9 genes were R2R3 type; 1 gene was R1R2R3 type; the rest were 1R type or MYB-related type, but the specific functional analysis has not been reported

Method used

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  • Peanut MYB transcription factor AhMYB31 and application thereof
  • Peanut MYB transcription factor AhMYB31 and application thereof
  • Peanut MYB transcription factor AhMYB31 and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0021] Embodiment 1: Acquisition of AhMYB31 gene

[0022] Select the relatively drought-resistant variety of peanut: Fenghua No. 1 (Lu Nongshenzi [2001] No. 017, provided by the medium-term bank of agricultural germplasm resources in Jiangsu Province), germinate and grow in the light incubator, and wait for the seedlings to grow to the 3-leaf stage, and carry out drought ( 20% PEG6000) and high salt (1% NaCl) treatment, after 24 hours of treatment, take part of the leaves and mix them in a mortar, quickly grind them into powder under liquid nitrogen, and use the RNA extraction kit (E.Z.N.A. TM PlantRNAKit (purchased from OMEGA Company) was used to extract the total RNA of the sample and digest it with DNAseI. The integrity of the total RNA was detected by 1% agarose gel electrophoresis, and the concentration and purity of the total RNA were detected by a NanoDrop-2000 spectrophotometer. The total RNA was reversed into cDNA by a reverse transcription kit.

[0023] According t...

Embodiment 2

[0027] Embodiment 2: Expression analysis of AhMYB31 gene

[0028] The peanut variety: Fenghua No. 1 was selected and germinated and grown in a light incubator. When the seedlings grew to the 3-leaf stage, they were subjected to drought (20% PEG6000) and high-salt (1% NaCl) treatments, respectively, and took 0h, 4h, 8h, The 12h leaf samples were frozen in liquid nitrogen and stored in a -80°C refrigerator. Then, the total RNA of the samples was respectively extracted with an RNA extraction kit (E.Z.N.A.TMPlantRNAKit, purchased from OMEGA Company), and reversed into cDNA.

[0029] Quantitative primers P3 and P4 (Table 1) were designed based on the AhMYB31 sequence, and the peanut 18SRNA gene primer (ChenNa, YangQingli, PanLijuan etal. Identification of 30MYBtranscriptionfactorgenesandanalysisoftheirexpressionduringabioticstressinpeanut (ArachishypogaeaL.). Gene, 2013, doi: 10.1016) was used as an internal reference peanut control (equivalent to It is constitutively expressed. I...

Embodiment 3

[0030] Example 3: Analysis of transcriptional self-activation activity of AhMYB31 gene

[0031] According to the AhMYB31 sequence, primers P5 and P6 with EcoR1 and BamH1 restriction sites were designed respectively (Table 1), and the target fragment was obtained by PCR amplification. Source recombinase (purchased from Shanghai Jierui Bioengineering Co., Ltd.) was transformed into Escherichia coli Trans1-T1 competent cells (purchased from Beijing Quanshijin Biotechnology Co., Ltd.), and the positive clones were sequenced and analyzed to ensure that the expression vector encoded The reading frame of the region was correct, thus the pGBKT7-AhMYB31 bait vector was obtained. Then the bait carrier plasmid was transformed into yeast Y2H competent (purchased from Shanghai Maiqi Biotechnology Co., Ltd.), and the transformation bacteria liquid was divided into 1, 10, 10 2 、10 3 Gradiently dilute, and then take 100 μL of the bacterial solution and spread the transformed bacterial solut...

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Abstract

The invention discloses a peanut R2R3-MYB transcription factor AhMYB31 and an application thereof, and belongs to the field of genetic engineering. The gene has a cDNA sequence of SEQ ID NO.1 and a coding amino acid sequence of SEQ ID NO.2. The AhMYB31 gene in peanuts is reported for the first time, fluorescence quantitative expression analysis indicates that the gene is subjected to high-salty inducible expression and functional verification with trans-arabidopsis, and results indicate that an over-expressed trans-genetic plant shows stronger salt resistance in comparison with a wild plant. Therefore, AhMYB31 can be introduced into a plant as a target gene, so that the salt resistance of the plant can be improved.

Description

technical field [0001] The invention discloses peanut MYB type transcription factor AhMYB31 and its application, belonging to the field of biogenetic engineering. Background technique [0002] The v-mybavian myeloblastosis viral oncogene homolog (MYB) transcription factor family is a family of transcription factors containing the MYB domain, and is a family with the largest number and the most diverse functions among plant transcription factors. The MYB domain is a peptide segment of about 51-52 amino acids, and there is a conserved tryptophan residue every 18-19 amino acids, which participate in the formation of the hydrophobic core in the spatial structure, and are important for maintaining the helix-turn- The helix (HTH: helix-turn-helix) configuration is of particular importance (Zhong et al., 2007). Paz-Ares et al. (1987) cloned the COLORED1 (C1) gene from corn grain aleurone for the first time, encoding a protein with a MYB domain, involved in anthocyanin synthesis. ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/415C12N15/29A01H5/00
CPCC07K14/415C12N15/8273
Inventor 刘永惠陈志德沈一
Owner JIANGSU ACADEMY OF AGRICULTURAL SCIENCES
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