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Strawberry salt stress related gene FvDIV and application thereof

A salt stress and gene technology, applied in the fields of molecular biology and genetic engineering, can solve the problem that the function of DIVARICATA gene is rarely reported, and achieve the effect of improving salt stress tolerance.

Inactive Publication Date: 2017-09-15
SHENYANG AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Another study showed that DIVARICATA (DIV), RADIALIS (RAD) and DICHOTOMA (DICH) were found to participate in the molecular regulation of bilaterally symmetrical flower development in the study of the model plant Antirrhinum (Wei Haichao et al., 2012), but the DIVARICATA gene The role in plant salt tolerance is rarely reported

Method used

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  • Strawberry salt stress related gene FvDIV and application thereof
  • Strawberry salt stress related gene FvDIV and application thereof
  • Strawberry salt stress related gene FvDIV and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0017] Embodiment 1, the cloning of strawberry salt stress gene FvDIV

[0018] (1) The diploid forest strawberry 'Ruegen' was used as the test material.

[0019] (2) RNA extraction: the modified CTAB method was used to extract the total RNA in the material, and then the first strand of cDNA was obtained by reverse transcription with RNA as a template using a reverse transcription kit.

[0020] (3) Cloning of the gene: using the first strand of reverse-transcribed fruit cDNA as a template, PCR amplification was performed using primers FvDIV-F and FvDIV-R, and the PCR product was recovered to obtain a 939bp target fragment.

[0021] FvDIV-F: 5'-AAAGGTACCATGTATAGGGGAATTGAAGTTC-3'

[0022] FvDIV-R: 5'-AAAGAATTCCTGATATTGCATCTCGAAACGC-3'

[0023] Among them, the first 3 bases at the 5' end of the primer are protective bases, the next 6 bases are enzyme cutting sites, and the first 9 bases at the 5' end are required for the construction of overexpression vectors and do not belong t...

example 2

[0024] Example 2, the construction of plant expression vector

[0025] (1) Using the plant expression vector pRI101-GFP-CaMV35S, select Kpn1 and EcoR1 (purchased from TaKaRa Company) to carry out double enzyme digestion on pRI101-GFP-CaMV35S and PMD18-T (purchased from TaKaRa Company) containing the target gene respectively; The large fragment of the vector and the small fragment of the target gene were ligated overnight with T4 DNA ligase and then transformed into Escherichia coli competent DH5a (purchased from Beijing Quanshijin Biotechnology Co., Ltd.).

[0026] (2) Select the correct recombinant plasmid verified by double enzyme digestion and send it to Jinweizhi Biotechnology Co., Ltd. for sequencing. Afterwards, Agrobacterium GV3101 competent cells were transformed, and the obtained Agrobacterium containing the recombinant plasmid was used to transform Arabidopsis.

example 3

[0027] Example 3. Verification of transgene function—Arabidopsis transformation screening and salt-tolerant phenotype analysis

[0028] 3.1 Arabidopsis transformation

[0029] When Arabidopsis (ecotype Columbia) inflorescences were formed, the tops of the inflorescences were cut off to induce the formation of side inflorescences, and the materials were watered thoroughly before transformation. Agrobacterium GV3101 positive clones containing the target gene FvDIV were selected. Inoculate in YEP liquid medium containing Kan (kanamycin) 50mg / L and Rif (rifampicin) 25mg / L, culture at 28°C, 200rpm shaking for 24h, take 1ml cultured bacteria liquid, add 50ml liquid YEP The liquid culture medium was placed at 28° C. and cultured with shaking at 200 rpm, so that OD600 = about 0.8.

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Abstract

The invention relates to strawberry salt stress related gene FvDIV and application thereof and belongs to the field of gene engineering in molecular biology. A full-length coding region sequence of the strawberry FvDIV for enhancing plant salt stress tolerance is shown as in SEQ ID NO: 1, and its amino acid sequence is shown as in SEQ ID NO: 2. The strawberry salt stress related gene FvDIV provided herein is also used to construct a plant expression vector of FvDIV gene, the constructed plant expression vector transforms Arabidopsis thaliana by means of Agrobacterium-mediated method, and the acquired transgenic plant has significantly improved salt stress tolerance. The strawberry salt stress related gene FvDIV and the application thereof provide theoretical basis and technical means for improving plant salt stress tolerance through gene engineering techniques, and are of great applicable value.

Description

technical field [0001] The invention belongs to the technical fields of molecular biology and genetic engineering, and in particular relates to a strawberry salt stress-related gene FvDIV and an application thereof. Background technique [0002] Strawberry is a perennial herb plant belonging to the Rosaceae Fragaria genus, a berry fruit tree (Lin Fengqi, 1986). Strawberry fruit is a false fruit, which develops from a receptacle (Lei Jiajun, 2006). Strawberry fruit has an attractive red color, soft and juicy taste, unique flavor, high nutritional value and medicinal value. It is known as the "Queen of Fruits" and is widely welcomed by the public (Wan Qinglin, 1994). [0003] The adaptability of plants to abiotic stress (drought, chilling damage, high salinity, etc.) has a very important impact on crop yield and quality, and is the bottleneck of agricultural development in many areas. Since the cultivation of strawberries in the northern region is carried out in solar greenh...

Claims

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Application Information

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IPC IPC(8): C12N15/29C12N15/82C12N15/11C07K14/415A01H5/00
Inventor 张俊祥石伟佳张志宏孙一平王保田
Owner SHENYANG AGRI UNIV
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