Abstract

Overexpression of the cytochrome P450 monooxygenase (cyp71av1) and cytochrome P450 reductase (cpr) genes increased artemisinin content in Artemisia annua (Asteraceae)

Author(s): Q. Shen Y.F. Chen T. Wang S.Y. Wu X. Lu L. Zhang F.Y. Zhang W.M. Jiang G.F. Wang K.X. Tang

Finding an efficient and affordable treatment against malaria is still a challenge for medicine. Artemisinin is an effective anti-malarial drug isolated from Artemisia annua. However, the artemisinin content of A. annua is very low. We used transgenic technology to increase the artemisinin content of A. annua by overexpressing cytochrome P450 monooxygenase (cyp71av1) and cytochrome P450 reductase (cpr) genes. CYP71AV1 is a key enzyme in the artemisinin biosynthesis pathway, while CPR is a redox partner for CYP71AV1. Eight independent transgenic A. annua plants were obtained through Agrobacterium tumefaciens-mediated transformation, which was confirmed by PCR and Southern blot analyses. The real-time qPCR results showed that the gene cyp71av1 was highly expressed at the transcriptional level in the transgenic A. annua plants. HPLC analysis showed that the artemisinin content was increased in a number of the transgenic plants, in which both cyp71av1 and cpr were overexpressed. In one of the transgenic A. annua plants, the artemisinin content was 38% higher than in the non-transgenic plants. We conclude that overexpressing key enzymes of the biosynthesis pathway is an effective means for increasing artemisinin content in plants. Finding an efficient and affordable treatment against malaria is still a challenge for medicine. Artemisinin is an effective anti-malarial drug isolated from Artemisia annua. However, the artemisinin content of A. annua is very low. We used transgenic technology to increase the artemisinin content of A. annua by overexpressing cytochrome P450 monooxygenase (cyp71av1) and cytochrome P450 reductase (cpr) genes. CYP71AV1 is a key enzyme in the artemisinin biosynthesis pathway, while CPR is a redox partner for CYP71AV1. Eight independent transgenic A. annua plants were obtained through Agrobacterium tumefaciens-mediated transformation, which was confirmed by PCR and Southern blot analyses. The real-time qPCR results showed that the gene cyp71av1 was highly expressed at the transcriptional level in the transgenic A. annua plants. HPLC analysis showed that the artemisinin content was increased in a number of the transgenic plants, in which both cyp71av1 and cpr were overexpressed. In one of the transgenic A. annua plants, the artemisinin content was 38% higher than in the non-transgenic plants. We conclude that overexpressing key enzymes of the biosynthesis pathway is an effective means for increasing artemisinin content in plants.

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