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Gene associated with synthesis of lutein and application

A technology of lutein and plant lutein, which is applied in the field of plant genetic engineering and can solve the problems that genes have not been isolated and reported

Inactive Publication Date: 2021-02-05
HEFEI UNIV OF TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

But for marigold, which is an important plant source of lutein, some important genes of its synthetic pathway have not been isolated and reported

Method used

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  • Gene associated with synthesis of lutein and application
  • Gene associated with synthesis of lutein and application
  • Gene associated with synthesis of lutein and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0015] Example 1: High-throughput transcriptome sequencing of marigold

[0016] Since the marigold genome has not been determined, in order to obtain the sequence of functional gene transcripts, a total of 9 samples were subjected to high-throughput DNA extraction by extracting RNA from leaves, immature flowers, and mature flowers of three single plants using tissue samples of marigold cultivars. Transcriptome sequence determination and assembly annotation.

[0017] 1. Reagents

[0018] The plant RNA extraction reagent TRIzol was purchased from Invitrogen Company, DNase I (Dnase I) was purchased from TaKaRa Company, the RNA library preparation kit (RNA Library Prep Kit) was from Beijing Biomike Biotechnology Co., Ltd., and the rest of the reagents were imported. Or domestic analytically pure products.

[0019] 2. Plant material

[0020] Marigold (Tagetes erecta L.) cultivar Juwang was bred by Chifeng Xinhui Gardening Company.

[0021] 3. Method

[0022] 3.1 RNA extraction...

Embodiment 2

[0055] Example 2: Cloning of Tagetes TeCHX Gene

[0056] According to the data assembly of Tagetes TeCHX gene in Example 1, use the primer design software Primer Premier to design PCR-specific primers, extract total RNA from mature flowers of Tagetes, and reverse transcription-PCR (reverse transcription-PCR, RT-PCR) clone to obtain TeCHX gene.

[0057] 1. Reagents

[0058] Plant RNA extraction reagent TRIzol was purchased from Invitrogen Company, DNase I (Dnase I) was purchased from TaKaRa Company, reverse transcriptase, dNTP, and high-fidelity DNA polymerase were purchased from Beijing Quanshijin Biotechnology Co., Ltd., and primers were purchased from Sangon Biotechnology Co., Ltd. Synthesized by Engineering (Shanghai) Co., Ltd., and the rest of the reagents were imported or domestically produced analytically pure products.

[0059] 2. Vector and strain

[0060] The cloning vector pEASY-Blunt Simple Cloning Vector was purchased from Beijing Quanshijin Biotechnology Co., L...

Embodiment 3

[0109] Embodiment 3: HPLC method measures the content of lutein in marigold flower

[0110] 1. Reagents

[0111] Standard lutein was purchased from Shanghai Aladdin Biochemical Technology Co., Ltd., 2,6-di-tert-butyl-4-methylphenol was purchased from Sinopharm Chemical Reagent Co., Ltd., and the rest of the reagents were imported or domestically produced. product.

[0112] 2. Method

[0113]Take samples of immature and mature marigold flowers, dry them at 45°C, grind the dried samples with liquid nitrogen, weigh 0.15 g, add 1 mL of ethanol (containing 0.1% 2,6-di-tert-butyl-4-methyl Phenol), vortex for 15s, add 400mL 50% KOH aqueous solution, vortex for 15s, place in 50°C water bath for 60min, vortex once every 15min. Extract 3 times with 3.3 mL of n-hexane, combine the extracts, and take 50 μL of the filtrate for loading. Chromatographic conditions: high performance liquid chromatography (WATERS, USA); column SymmetryC18 (250mm×4.6mm, 5μm); mobile phase: acetonitrile: dic...

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Abstract

The invention discloses a gene associated with synthesis of lutein and an application. The gene TeCHX has a nucleotide sequence as shown in SEQ ID NO.1, and an amino acid sequence as shown in SEQ ID NO.2. According to the invention, the TeCHX gene is obtained from marigold through cloning. Real-time quantitative analysis results show that the expression level of TeCHX in mature marigold flowers with significantly increased lutein content is significantly higher than that in immature marigold flowers. When TeCHX is expressed in tobacco, the lutein content of the tobacco can be increased. The gene as shown in SEQ ID NO.1 plays a role in biosynthesis of lutein. The carotene hydroxylase gene TeCHX cloned from pigment marigold can be used for increasing the lutein content of plants.

Description

technical field [0001] The invention relates to the technical field of plant genetic engineering, in particular to a gene cloned from marigold and the application of the gene in plant lutein synthesis. Background technique [0002] Carotenoids are an important class of pigment components in plants. They not only play a role in photosynthesis, but are also used by plants for adversity protection, attracting pollinators, and for the synthesis and metabolism of other regulatory substances. Marigold (Tagetes erecta L.), an annual herbaceous plant of the Compositae family native to Mexico and the Americas, has become an ornamental plant because of its bright color, various flower colors from yellow to orange red, and its long flowering period and easy cultivation. The main pigment components in marigold flowers are carotenoids, and the content is relatively high, so this ornamental plant has also become a major plant resource for extracting carotenoids, especially lutein. [000...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/53C12N9/02C12N15/82A01H5/02A01H6/82
CPCC12N9/0073C12N15/825C12Y114/13129
Inventor 牛向丽马飞张恩启徐焕焕云俊文谭永毅聂婧孙甲玲张芳毓
Owner HEFEI UNIV OF TECH
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