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A myrothamnus flabellifolia gene MfWRKY17 and application thereof

A kind of Mirosa, gene technology, applied in the direction of application, genetic engineering, plant genetic improvement, etc., can solve the problem that the stress-resistant WRKY transcription factor, gene and its molecular mechanism are not yet known in Mirosa.

Inactive Publication Date: 2019-01-11
SICHUAN AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, in the past, researchers have mainly studied the drought resistance mechanism of the plant through morphological, physiological, biochemical and other aspects, but the genes and molecular mechanisms related to stress resistance are still unclear, and the stress-resistant WRKY transcription factor in Miroma has not yet been found. Related reports on its application

Method used

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  • A myrothamnus flabellifolia gene MfWRKY17 and application thereof
  • A myrothamnus flabellifolia gene MfWRKY17 and application thereof
  • A myrothamnus flabellifolia gene MfWRKY17 and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0032] Example 1 Cloning of Miluo MfWRKY17 gene

[0033] 1. Take the leaves of Miluo wood, quick-frozen them in liquid nitrogen, and store them in the refrigerator at -80℃ for total RNA extraction; the total RNA was extracted using the Plant Total RNA Isolation Kit purchased by Chengdu Lanbo Biological Company; Miluo cDNA For the synthesis of Reverse Transcriptase M-MLV (RNaseH-) from Dalian Bao Biotechnology Company, the first-strand synthesis was performed according to its product instructions.

[0034] Using the first strand of cDNA synthesized by the above kit as the amplification template, the designed F: 5'-TCCCCCGGGATGGCGGTTGACTTGCTC-3' (SEQ ID NO. 3) and R: 5'-GACTAGTTCATGGTGTTGACTCAAACA-3' (SEQ ID NO. 4 ) Are primers. For subsequent enzyme digestion and recombination and ligation, two enzyme sites, SmaI and SpeI, are added when designing primers, and cDNA is amplified by PCR. The amplification system is shown in Table 1, and the amplification conditions are: 95 Pre-denatu...

Embodiment 2

[0038] Example 2 Sequence homology and homology analysis of MfWRKY17

[0039] According to the results of sequence sequencing, the sequence alignment 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 factor. The transcription factor was compared with the protein sequence of other WRKY transcription factor family members and found that it The zinc finger structure type is C2H2 type. According to the structural characteristics of the DNA binding domain, the MfWRKY17 protein belongs to the second type of WRKY transcription factor family; further use the aligned amino acid sequences of species with higher homology to construct a phylogenetic tree (such as figure 2 ), it is found that MfWRKY17 has a long relationship with other species, and its domain is relatively conservative.

Embodiment 3

[0040] Example 3 Expression levels of MfWRKY17 in different dehydration periods of Miluo

[0041] The Trizol method was used to extract the total RNA from the leaves of Miluo at 0, 10%, and 25% dehydration stage. Using the extracted total RNA as a template, the reverse transcription and reverse transcription were performed using the Reverse Transcriptase M-MLV (RNaseH-) of Dalian Bao Biotechnology Company. Obtain cDNA.

[0042] Among them, Actin is used as an internal control, MfWRKY17 and Actin primers are designed as follows,

[0043] Actin primer:

[0044] F: 5'-ATCACCATTGGGGCTGAACG-3' (SEQ ID NO.5)

[0045] R: 5'-GTCGACCCACCACTAAGGAC-3' (SEQ ID NO. 6)

[0046] MfWRKR17 primer:

[0047] F: 5'-TCCTCAATCGCAGACGTTCA-3' (SEQ ID NO. 7)

[0048] R: 5'-CTGCGATATGCTAACGCTCTC-3' (SEQ ID NO. 8)

[0049] Carry out RT-PCR reaction based on the above primers. Each reaction has 3 technical replicates. The test results are as follows image 3 Show from image 3 It can be seen that the MfWRKY17 gene of...

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Abstract

The invention discloses a myrothamnus flabellifolia gene MfWRKY17 and an application thereof. The nucleotide sequence of the gene is shown in SEQ ID NO. 1; The amino acid sequence of the protein encoded by the gene is shown in SEQ ID NO: 2. The invention improves the stress resistance performance of the plant and improves the stress resistance performance of the plant through the research on the drought resistance WRKY transcription factor of Milo.

Description

Technical field [0001] The invention belongs to the technical field of plant genetic engineering, and specifically relates to a milo gene MfWRKY17 and its application. Background technique [0002] Drought and salinization are one of the main non-biological disasters facing plants today. The main mechanism of action is to make the internal and external osmotic pressure of the plant inconsistent and cause dehydration of the body. The population growth and the rapid development of urban construction have not only led to increasingly fierce competition for civil, industrial, and agricultural water use, but also increased pollution, damage to the ecosystem, water shortages, and increasing soil salinization. Therefore, discovering stress resistance genes and cultivating new plant varieties with strong stress resistance is important for accelerating the construction of a resource-saving and environment-friendly society, vigorously developing a green economy, enhancing sustainable devel...

Claims

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Application Information

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IPC IPC(8): C12N15/29C12N15/82C07K14/415A01H5/00A01H6/20
Inventor 黄卓蒋才忠郭涵度刘玲靳斯涵朱培蕾彭玉婷李西马均蔡仕珍邱嘉睿陈佳向香盈
Owner SICHUAN AGRI UNIV
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