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A drought-resistant transcription factor pbrerf109 and its preparation method, application, encoded protein and application

A transcription factor and protein technology, applied in the field of plant genetic engineering, can solve the problem of drought resistance of unreported ERF transcription factors

Active Publication Date: 2020-10-23
NANJING AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there is no report in the prior art that ERF transcription factors can resist drought

Method used

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  • A drought-resistant transcription factor pbrerf109 and its preparation method, application, encoded protein and application
  • A drought-resistant transcription factor pbrerf109 and its preparation method, application, encoded protein and application
  • A drought-resistant transcription factor pbrerf109 and its preparation method, application, encoded protein and application

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preparation example Construction

[0038] In the present invention, the preparation method of the drought-resistant transcription factor PbrERF109 comprises the following steps:

[0039] Using pear cDNA as a template, the drought-resistant transcription factor PbrERF109 was obtained by PCR amplification with transcription factor primers;

[0040] The transcription factor primer pair includes a transcription factor upstream primer and a transcription factor downstream primer, the transcription factor upstream primer has the nucleotide sequence shown in SEQ ID No.3, and the transcription factor downstream primer has the nucleotide sequence shown in SEQ ID No.4 The nucleotide sequence shown.

[0041] In the present invention, the method for preparing pear cDNA preferably comprises: extracting RNA from leaves of Pear pear, and reverse-transcribing the obtained RNA to obtain cDNA. The method for extracting RNA is not particularly limited in the present invention, and the method for extracting plant RNA conventional...

Embodiment 1

[0066] Cloning and expression analysis of PbrERF109 gene

[0067] RNA was extracted from the leaves of Duli pear under drought and dehydration treatment, and cDNA was obtained by reverse transcription. In the present invention, after obtaining the cNDA, the obtained cDNA is used as a template, and a specific transcription factor primer pair is used to obtain the drought-resistant transcription factor PbrERF109, the specific sequence of which is shown in SEQ ID No.1.

[0068] For RNA extraction, Plant Total RNA Isolation Kit Plus (Foregene, RE-05022) was used, and the operation instructions provided by the kit were followed. First-strand cDNA was synthesized using First Script Strand cDNASynthesis SuperMix (Transgene, AE301-02) reverse transcription kit (operated according to the instructions provided by the kit).

[0069] The primer pair for amplifying the drought-resistant transcription factor PbrERF109 is: Forward primer: PbrERF109 Forward, 5'-GA AGATCT TATGCCCTTCCATGCGAA...

Embodiment 2

[0079] Analysis of subcellular localization and transcriptional activation of PbrERF109 gene

[0080] Since the PbrERF109 gene is a transcription factor, the present invention uses tobacco mesophyll cells to study the subcellular localization of the PbrERF109 gene. The entire ORF reading frame of the PbrERF109 gene was amplified by RT-PCR, and two restriction sites, XbaI and BamHI, were added at both ends of the amplification primer. The amplification primer is (forward primer PbrERF109-F2: 5'- TCTAGA ATGCCCTTCCATGCGAATCGGATA-3'(SEQ ID No.11); reverse primer PbrERF109-R2: 5'- GGATCCCCTAGTTCGTGGATAACCA-3' (SEQ ID No.12)), firstly digest the amplified product. In the present invention, the temperature for the double digestion of the pear drought-induced transcription factor PbrERF109 is 37° C., and the time for the double digestion is 12 hours; the total volume of the double digestion system is 20 μl, including the PCR purified product of the pear drought-induced transcripti...

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Abstract

The invention provides a drought-resistant transcription factor PbrERF109, a preparation method and application of the transcription factor PbrERF109, encoded protein and application of the encoded protein, and belongs to the technical field of plant genetic engineering. The drought-resistant transcription factor PbrERF109 has a nucleotide sequence shown as ESQ ID No.1. Compared with a wild strain, the drought resistance of a transgenic overexpression strain obtained by transferring the drought-resistant transcription factor PbrERF109 into tobacco and pyrus ussuriensis is greatly improved, thecontents of hydrogen peroxide and malondialdehyde in the transgenic overexpression strain of tobacco are both lower than those of the wild strain, the residual amount of active oxygen in the plant body is lower, the cell damage is smaller, and thus the drought resistance of tobacco and pyrus ussuriensis is improved.

Description

technical field [0001] The invention belongs to the field of plant genetic engineering, and specifically relates to a drought-resistant transcription factor PbrERF109, its preparation method, application, encoded protein and application. Background technique [0002] Plants often encounter some adversity stresses during their growth cycle, including biotic stresses and abiotic stresses. Abiotic stresses mainly include drought, high temperature, low temperature, heavy metal stress, etc. Among them, drought affects the growth and development of plants, Yield quality and one of the important factors limiting the geographical distribution of plants. It is now clear that the survival of plants under drought stress mainly depends on the metabolism in the body and the adaptive changes in plant morphology, including a series of physiological and biochemical reactions to establish a systemic defense state and respond to drought stress at multiple levels , the expression of many drou...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/29C12N15/10C07K14/415C12N15/84A01H5/00A01H6/82A01H6/74
CPCC07K14/415C12N15/1096C12N15/8273C12Q2531/113
Inventor 张绍铃胡轼黄小三赵梁怡邢才华董慧珍高俊芝刘月李凌陶书田
Owner NANJING AGRICULTURAL UNIVERSITY
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