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Method for improving content of artemisinin in sweet wormwood herb through transferring F3H gene

A kind of artemisinin and transgenic technology, which is applied in horticultural methods, genetic engineering, plant gene improvement, etc., can solve the problems of artificial synthesis, such as difficulty, feasibility, and inability to fully meet market demand, and achieve the effect of stabilizing new drug sources

Inactive Publication Date: 2016-08-17
SHANGHAI JIAO TONG UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0003] However, the content of artemisinin in the plant Artemisia annua is very low, large-scale commercial production is limited, and it cannot fully meet the global market demand
Due to the complex structure of artemisinin, the artificial synthesis is difficult, the yield is low, and the cost is high, so it is not feasible
Production of artemisinin through yeast engineering, the initial investment cost is high, and the limited output cannot meet the demand

Method used

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  • Method for improving content of artemisinin in sweet wormwood herb through transferring F3H gene
  • Method for improving content of artemisinin in sweet wormwood herb through transferring F3H gene
  • Method for improving content of artemisinin in sweet wormwood herb through transferring F3H gene

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Embodiment

[0031] Step 1, cloning of Artemisia annua F3H gene

[0032] (1) Extraction of total RNA from Artemisia annua genome

[0033]The total RNA of Artemisia annua genome was extracted using RNA kit from TIANGEN Company. Take 50-100 mg young leaves of Artemisia annua and quickly grind them into powder in liquid nitrogen, add 550 μL of lysis solution RL (mercaptoethanol has been added), and vortex vigorously to mix. Centrifuge at 9800rpm for 5min, transfer 450μL to filter column CS, place CS in a collection tube, centrifuge at 12000rpm for 5 minutes, carefully draw 400μL of the supernatant in the collection tube into an RNase-free centrifuge tube, slowly add 0.5 times the volume of the supernatant mixed with absolute ethanol, transferred to the adsorption column CR3, centrifuged at 12,000 rpm for 1 min, discarded the waste liquid, and put CR3 back into the collection tube. Add 350 μL protein-removing solution RW1 to CR3, centrifuge at 12,000 rpm for 1 min, discard the waste liquid, ...

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Abstract

The invention relates to a method for improving the content of artemisinin in sweet wormwood herb through transferring F3H gene. The method comprises the following steps: cloning flavanone-3-hydroxylase (F3H) gene in sweet wormwood herb to construct an F3H gene-containing plant expression vector, mediating with Agrobacterium tumefaciens, transferring the F3H gene to the sweet wormwood herb, and regenerating a plant, carrying out PCR detection on the integration condition of the exogenous target gene F3H, carrying out HPLC-ELSD detection on the content of artemisinin in transgenic sweet wormwood herb, and screening to obtain transgenic sweet wormwood herb plants with improved content of artemisinin. The content of artemisinin in the obtained transgenic sweet wormwood herb is substantially improved, and the highest content of artemisinin is 1.95 times the content of artemisinin in a nontransgenic control plant, so the method for improving content of artemisinin in sweet wormwood herb lays a foundation for large-scale production of artemisinin through using the transgenic sweet wormwood herb.

Description

technical field [0001] The invention relates to a method for increasing the content of artemisinin in the field of biotechnology, in particular to a method for increasing the content of artemisinin in Artemisia annua by transfecting F3H gene. Background technique [0002] Artemisia annua L. is an annual herb belonging to the family Asteraceae. Artemisinin, a sesquiterpene lactone oxide containing a peroxy bridge extracted from its aerial part, is currently the most widely used and effective antimalarial drug, especially for cerebral malaria and anti-chloroquine malaria. . Currently, artemisinin combination therapies (ACTs) are the most effective treatment for malaria recommended by the World Health Organization. With the gradual deepening of pharmacological research on artemisinin, scientists have discovered that artemisinin and its derivatives also have anti-inflammatory, anti-tumor, anti-cancer and immunomodulatory functions. [0003] However, the content of artemisinin...

Claims

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Application Information

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IPC IPC(8): C12N15/84C12N9/02C12N15/53A01H5/00A01H4/00
CPCA01H4/00C12N9/0071C12N15/8205C12N15/8243C12Y114/11009
Inventor 赵静雅王路尧张颖付雪晴张婷婷张利达钱虹妹唐克轩
Owner SHANGHAI JIAO TONG UNIV
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