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Construction and transformation of chamomile farnesyl diphosphate synthase (FPS) gene eukaryotic expression vector

A technology of nicyl pyrophosphate synthase and eukaryotic expression vector, which is applied in the direction of genetic engineering, plant gene improvement, and the introduction of foreign genetic material using vectors, etc., to achieve the effect of increasing production and making great contributions

Inactive Publication Date: 2012-12-12
ANHUI AGRICULTURAL UNIVERSITY
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Problems solved by technology

However, there is no research on the enzymes and genes in the volatile oil of chamomile that control the metabolic pathway of matricaria azulene

Method used

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  • Construction and transformation of chamomile farnesyl diphosphate synthase (FPS) gene eukaryotic expression vector
  • Construction and transformation of chamomile farnesyl diphosphate synthase (FPS) gene eukaryotic expression vector
  • Construction and transformation of chamomile farnesyl diphosphate synthase (FPS) gene eukaryotic expression vector

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Embodiment Construction

[0048] 1. Materials and methods

[0049] 1.1 Experimental materials

[0050] 1.1.1 Plant materials: chamomile flowers (collected from the Nongcui Garden of Anhui Agricultural University), aseptic seedlings of Yunyan 85 tobacco (provided by the tissue culture room of the College of Life Sciences, Anhui Agricultural University).

[0051] 1.1.2 Strains and plasmids: Escherichia coli DH5a competent cells were purchased from TAKARA Company, Agrobacterium EHA105 and pBI121 vectors were donated by Mr. Jiang Haiyang, School of Life Sciences, Anhui Agricultural University, pEASY-T4 Zero Cloning Vector and Trans1-T1 competent cells were purchased From Beijing Quanshijin Biotechnology Co., Ltd.

[0052] 1.1.3 Main biological reagents: Restriction enzymes SacI, BamHI, T4DNA ligase, rtaq were purchased from TAKARA company, TransStart Taq DNA Polymerase was purchased from Beijing Quanshijin Biotechnology Co., Ltd., RNAiso plus, DNA molecular weight standard, DNA gel recovery kit and plasmid...

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Abstract

The invention discloses a construction and transformation method of a chamomile farnesyl diphosphate synthase (FPS) gene eukaryotic expression vector, comprising the steps of: extracting an RNA (ribonucleic acid) from chamomile petals, and inversely transcribing the RNA into cDNA (complementary deoxyribonucleic acid) with total chamomile RNAs as a template; carrying out PCR (Polymerase Chain Reaction) amplification using a cDNA as a template to obtain a target fragment; connecting the target fragment with a T4 ligase; transforming a connection product to an escherichia coli DH5 alpha competent cell to obtain an eukaryotic expression vector PBI121 / FPS; then introducing a recombinant expression vector to agrobacterium tumefaciens EHA105; and infecting the plant Yunyan85 by using the agrobacterium tumefaciens EHA105 to finally obtain a resistant tobacco plant. An FPS gene is cloned by using a gene engineering technology, the callus tissues of the plant are infected by virtue of the agrobacterium tumefaciens, and the construction and transformation method lays a foundation for improvement of chamazulene yields through expression of the FPS gene in a plant frond.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a chamomile farnesyl pyrophosphate synthase gene cloning, eukaryotic expression and application. Background technique [0002] Chamomile (Matricaria chamomilla L.) is an annual herb of the genus Matricaria in the Asteraceae family. The whole plant is fragrant and contains aromatic volatile oil. Chamomile is anti-inflammatory, anti-fungal, antispasmodic [1-3] , Improve the symptoms of lung cancer patients and so on. Chamomile originates in Europe and was introduced into my country at the beginning of this century. It is rarely cultivated in Beijing, Shanghai, Jiangsu, Hunan and other places. Chamomile contains a variety of chemical components: volatile oil, flavonoid glycosides, guaiazolactones, coumarin, polysaccharides, organic acids and other components. Chamomile volatile oil is the most important active ingredient in inflorescences. Its volatile oil is widely used in food, bev...

Claims

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Application Information

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IPC IPC(8): C12N15/66C12N15/54C12N15/82
Inventor 袁艺杨秀梅邰玉玲
Owner ANHUI AGRICULTURAL UNIVERSITY
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