Application of soybean transcription factor GmWRKY23 gene in stress resistance

A transcription factor and transgenic technology, applied in the field of genetic engineering and botany, to achieve obvious antibacterial effect

Active Publication Date: 2018-09-28
黑龙江省农业科学院大豆研究所
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

A large number of WRKY genes have been predicted, but few soybean genes have been cloned for functional identification. With the development of genome-wide analysis, more and more WRKY proteins have been found in many plant species

Method used

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  • Application of soybean transcription factor GmWRKY23 gene in stress resistance
  • Application of soybean transcription factor GmWRKY23 gene in stress resistance
  • Application of soybean transcription factor GmWRKY23 gene in stress resistance

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0051] Example 1 Expression pattern of GmWRKY23 in young roots of different resistant cultivars under Pythium infection

[0052] Soybean germplasm Huibuzhidou (Hb-2), Heihe 38 (Hh38), and Suinong 29 (sh29) were provided by the Soybean Research Institute of the Heilongjiang Academy of Agricultural Sciences; Pythium was provided by the Soybean Research Institute of the Heilongjiang Academy of Agricultural Sciences.

[0053] Ⅰ. Soybean germination test

[0054] Wash the soybean seeds of Hb-2, Hh38 and sh29 with distilled water, dry them, and sterilize them with chlorine gas for 12-16 hours; then soak them in sterile deionized water for 12 hours, and place them evenly in a container with WA medium (water agar medium) on the plate, about 2cm away from the edge of the plate, placed in a growth chamber at room temperature for germination culture.

[0055] Ⅱ. Culture of Bacteria

[0056] Using a sterilized hole puncher, from the V8 medium for cultivating Pythium (V-8 fruit juice 200...

Embodiment 2

[0064] Example 2 The expression pattern of GmWRKY23 gene under the infection of different root rot pathogens

[0065] Rhizoctonia solani was provided by the Soybean Research Institute of Heilongjiang Academy of Agricultural Sciences; Phytophthora was provided by the Plant Protection Office of Hejiang Branch of Heilongjiang Academy of Agricultural Sciences.

[0066] The disease-resistant variety Hb-2 was used as a test material to explore the gene expression profiles of Phytophthora, Pythium, and Rhizoctonia solani infecting young soybean roots. The specific test methods refer to Example 1. The result is as Figure 2-4 , where TPM (Transcript per million clean Tags) is the number of copies of the transcript per million clean Tags. The expression level of GmWRKY23 gene was the highest after 6 hours of Pythium infection, and the expression level decreased at 24 hours; under the infection of Phytophthora, the expression level of GmWRKY23 in soybean young roots was the highest at ...

Embodiment 3

[0067] Cloning of embodiment 3 GmWRKY23 gene

[0068] Ⅰ. Extraction of RNA

[0069] The RNAprep pure plant total RNA extraction kit from TIANGEN Company was used to extract the total RNA of the hypocotyls of the soybean variety Hb-2 inoculated with Pythium, and the purity of the extracted total RNA was tested.

[0070] (1) The tweezers, medicine spoon and mortar used in the test were first grilled with alcohol for 15 minutes. The tips and centrifuge tubes for RNA extraction are RNase-Free products from Axygen Company.

[0071] (2) Weigh 100 mg of the hypocotyl tissue of soybean variety Hb-2 6 hours after inoculation with Pythium, quickly grind it into powder in liquid nitrogen, add 500 μL SL (check whether β-mercaptoethanol has been added before use), and immediately vortex Shake vigorously to mix; centrifuge at 12,000rpm (~13,400×g) for 2min.

[0072] (3) Transfer the supernatant to the filter column CS (the filter column CS is placed in the collection tube), centrifuge at...

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Abstract

The invention discloses the application of a pythium-induced soybean transcription factor GmWRKY23 gene in stress resistance. The application comprises the following specific steps: infecting a soybean by pythium to obtain a soybean with high expression of the GmWRKY23 gene; constructing a transgenic plant expression vector; converting the plant expression vector into agrobacterium to obtain recombinant plasmid-containing GV3101; infecting a cotyledonary node of an ungerminated soybean JK506 by the recombinant plasmid-containing GV3101, screening harvested T0 soybean seeds by using an MS culture medium containing 5 to 15 mg/L of glufosinate-ammonium, planting a generation T1, and screening T1 to obtain generation-T2 seeds for phenotypic identification test, resistance identification of transgenic soybean plants, in-vitro identification of transgenic soybean pathogen resistance and salt-tolerant experiment of transgenic soybeans. The transgenic plant of the invention has obvious bacteriostatic action on fusarium oxysporum; the mature period of the transgenic plant is earlier than that of a receptor contrast; the transgenic plant has salt tolerance.

Description

technical field [0001] The invention belongs to the fields of genetic engineering and botany, and in particular relates to the application of soybean transcription factor GmWRKY23 gene in stress resistance. Background technique [0002] Transcription factors are a class of proteins that can directly bind or act indirectly on the gene promoter region, and can activate or inhibit gene transcription. WRKY gene is a unique type of zinc finger transcription factor in plants, named for its conserved WRKYGOK domain. WRKY proteins can be divided into three categories (Groups-I-III). The WRKYGOK sequence has been reported in many organisms one after another. The studies on Arabidopsis, rice, maize and wheat all show that the WRKY gene is a large family composed of multiple members. Plant WRKY transcription factor genes are widely involved in the response to biotic and abiotic stresses. Under stress, plants can sense and transmit signals, activate downstream transcription factors, st...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/82C12N15/29A01H5/00A01H6/54
CPCC12N15/8273C12N15/8282
Inventor 魏崃刘丽君王伟威
Owner 黑龙江省农业科学院大豆研究所
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