Tamarix hispida MYB transcription factor coding gene and application thereof

A technology of tamarisk bristles and transcription factors, which can be applied in the fields of application, genetic engineering, and plant genetic improvement, and can solve problems such as inability to deeply understand the mechanism of salt tolerance

Active Publication Date: 2018-07-31
NORTHEAST FORESTRY UNIVERSITY
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Some studies only show that the expression of the MYB gene is induced by salt stress and may be involved in the regulation of salt tolerance in plants, but the mechanism of salt

Method used

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  • Tamarix hispida MYB transcription factor coding gene and application thereof
  • Tamarix hispida MYB transcription factor coding gene and application thereof
  • Tamarix hispida MYB transcription factor coding gene and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035] Example 1: Cloning, bioinformatics analysis and domain verification of the ThMYB gene of Tamarix bristles

[0036] 1. Cloning of the ThMYB gene of Tamarix bristles

[0037] 1.1 Extraction of total RNA from Tamarix brix

[0038] Seeds of Tamarix briatus were sown in artificial soil with a ratio of carbon soil to sand of 2:1, placed in a relative humidity of 65–75%, and a light intensity of 400 μmol m -2 ·s -1 , the average temperature is 22 ± 2 ° C greenhouse growth. After 2 months of growth, use 0.3mol·L -1 NaHCO 3 The roots were irrigated with the solution for stress treatment. After 0h (before stress treatment), 12h, 24h and 48h of treatment, the leaves and roots were taken and put into liquid nitrogen for quick freezing for RNA extraction. The CTAB method was used to extract RNA in each treatment period, and the extracted RNA samples were sent to Shenzhen Huada Gene Technology Co., Ltd. for the construction and sequencing of the transcriptome library.

[0039] ...

Embodiment 2

[0058] Embodiment 2: Construct the overexpression vector of Tamarix bristly ThMYB gene

[0059] According to the multiple cloning site of the plant overexpression vector pROKII and the characteristics of the ThMYB gene, the XbaI and KpnI restriction endonuclease sites were introduced into the 5' end and the 3' end of the ThMYB gene respectively, such as SEQ ID NO: 9 and SEQ ID The gene-specific upstream and downstream primers pROKⅡ-ThMYB-F and pROKⅡ-ThMYB-R indicated by NO: 10 were used to clone the ThMYB gene by RT-PCR using Tamarix cDNA as a template.

[0060] The RT-PCR reaction system was 20 μL, including 2 μL template, 1 μL each of pROKⅡ-ThMYB-F and pROKⅡ-ThMYB-R specific primers (10 μmol / L), 0.4 μL dNTP Mix (10 mmol / L), 10× LA Taq PCR buffer 2.0μL, LA Taq (5U / μL) 0.25μL. The reaction program was pre-denaturation at 94°C for 3 min; denaturation at 94°C for 30 s, annealing at 58°C for 30 s, extension at 72°C for 1 min, and 35 cycles; extension at 72°C for 7 min.

[0061]...

Embodiment 3

[0068] Example 3: Transient transformation of Tamarix bristles ThMYB gene

[0069] In this example, the overexpression vector pROKII-ThMYB (OE) constructed in Example 2 and the suppressed expression vector pFGC5941-ThMYB (SE) constructed in Comparative Example 1 were transformed into Tamarix bristles by using the transient transformation technique, so that the ThMYB gene can be transiently overexpress or suppress expression. At the same time, the empty pROKII vector was transformed into Tamarix brix as a control (CON).

[0070] The specific method is as follows:

[0071] 1. Select Tamarix bristles tissue-cultured seedlings subcultured for 30-35 days, with a plant height of 2.5-3.0 cm, and put the selected Tamarix bristles tissue-cultured seedlings into the high-osmotic treatment solution to soak for 5-10 minutes;

[0072] 2. Rapidly transfer the T. tumefaciens tissue culture seedlings treated with hypertonicity in step 1 to the infection solution containing the Agrobacterium t...

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Abstract

The invention belongs to the technical field of plant genetic engineering breeding, and particularly relates to a tamarix hispida MYB transcription factor coding gene and application thereof. The tamarix hispida ThMYB gene provided by the invention can be used for improving salt resistance of plants or breeding salt-resistant transgenic plants. Overexpression and suppression expression vectors ofthe ThMYB gene are respectively constructed, transgenic tamarix hispida is obtained by transient infection, histochemical staining and physiological index measurement prove that an overexpression strain of the ThMYB gene has obvious salt resistance; when the ThMYB gene is transferred into arabidopsis, after transgenic T3-generation arabidopsis seeds are stressed by 150mM NaCl, an average germination rate of transgenic T3-generation arabidopsis seeds is 6.21 times of that of wild arabidopsis seeds, root lengths of the transgenic T3-generation arabidopsis seeds are 1.41 times of that of the wildtransgenic T3-generation arabidopsis seeds, and a fresh weight of the transgenic T3-generation arabidopsis seeds is 1.66 times of that of the wild transgenic T3-generation arabidopsis seeds. Both homologous and heterogenous overexpression results of the ThMYB gene show that the gene can obviously improve salt resistance of the transgenic plants and is an excellent gene for salt-resistant geneticengineering breeding.

Description

technical field [0001] The invention belongs to the technical field of plant genetic engineering and breeding, and in particular relates to a gene encoding tamarisk bristles MYB transcription factor and an application thereof. Background technique [0002] Plant adversity stress includes biotic stress and abiotic stress. Biological stresses are mainly diseases and insect pests, weeds, etc.; abiotic stresses are mainly drought, high salinity, waterlogging, high temperature and freezing damage. Under stress stimulation, some transcription factors are overexpressed to transmit and amplify the signal to regulate the expression of corresponding downstream functional genes, thereby improving the stress resistance of plants. Since transcription factors can regulate the expression of multiple downstream genes, in plant genetic engineering breeding, compared with the transfer of a functional gene, the transfer of a transcription factor can improve the stress resistance of plants. ...

Claims

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Application Information

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IPC IPC(8): C12N15/29C12N15/82C07K14/415A01H5/00A01H6/20A01H6/82
CPCC07K14/415C12N15/8273
Inventor 高彩球张腾倩刘中原李新苹吕佳欣雷晓锦
Owner NORTHEAST FORESTRY UNIVERSITY
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