The role of licorice DXS knockout and overexpression in glycyrrhizic acid biosynthesis
10.16438/j.0513-4870.2021-0225
- VernacularTitle:甘草DXS基因过表达及表达沉默对甘草酸生物合成的影响研究
- Author:
Lin YANG
1
;
Dou-dou WANG
1
;
Shao-kai TIAN
1
;
Zhi-xin ZHANG
1
;
Jia-ming HOU
1
;
Yao XIAO
2
;
Ying LIU
1
Author Information
1. School of Life Sciences, Beijing University of Chinese Medicine, Beijing 102488, China
2. School of Chinese Pharmacy, Beijing University of Chinese Medicine, Beijing 102488, China
- Publication Type:Research Article
- Keywords:
glycyrrhizic acid;
1-deoxy-D-xylulose-5-phosphate synthase;
gene overexpression;
CRISPR/Cas9;
hairy root
- From:
Acta Pharmaceutica Sinica
2021;56(7):2025-2032
- CountryChina
- Language:Chinese
-
Abstract:
1-Deoxy-D-xylulose-5-phosphate synthase (DXS) is a rate-limiting enzyme involved in the 2-C-methyl-D-erythritol 4-phosphate (MEP) pathway for terpenoid precursor biosynthesis. DXS plays an essential role in glycyrrhizic acid (GA) biosynthesis. Based on our previous transcriptome study, there was a negative correlation between DXS expression and GA content. Therefore, we explored the regulatory role of DXS in GA biosynthesis using both gene overexpression and gene knockout in a hairy root culture system. DXS was cloned from Glycyrrhiza glabra L. (GenBank Accession No. MN158121). A plant binary expression vector pCA-DXS was constructed by a gene fusion method. The sgRNA sequence was designed based on the first exon of DXS to construct the gene editing vector pHSE-DXS. Hairy roots overexpressing or knocking out DXS were generated through an Agrobacterium-mediated method with licorice hypocotyls as explants. Wild-type hairy roots and negative control hairy roots containing empty plasmids were also evaluated. UPLC was used to determine the GA content in each licorice hairy root line. Results showed that the content of GA in the hairy root group knocking out DXS was significantly higher than that in the wild-type and negative control groups, while in the hairy root group overexpressing DXS was significantly lower, suggesting that DXS plays a negative role in GA biosynthesis. This study provides a foundation for determining the function of DXS in terpenoid metabolism and for further establishment of a molecular regulatory network of GA biosynthesis.