Cloning and Expression Analysis of Squalene Epoxidase Gene in Poria cocos
10.13422/j.cnki.syfjx.20230111
- VernacularTitle:茯苓鲨烯环氧酶基因克隆与表达分析
- Author:
Xiaoliu LIU
1
;
Zhenni XIE
2
;
Can ZHONG
2
;
Jing XIE
2
;
Shuihan ZHANG
1
;
Jian JIN
1
Author Information
1. Graduate School， Hunan University of Chinese Medicine， Changsha 410208， China
2. Institute of Chinese Medicine Resources， Hunan Academy of Chinese Medicine， Changsha 410013， China
- Publication Type:Journal Article
- Keywords:
Poria cocos;
squalene epoxidase （SE）gene;
gene cloning;
bioinformatics analysis;
gene expression
- From:
Chinese Journal of Experimental Traditional Medical Formulae
2023;29(4):144-152
- CountryChina
- Language:Chinese
-
Abstract:
ObjectiveTo clone squalene epoxidase （SE）， a potential key rate-limiting enzyme involved in the synthesis pathway of Poria cocos triterpenes， from P. cocos and analyze for bioinformatics and expression. MethodThe total RNA was extracted by the kit and reverse-transcribed to cDNA. Specific primers were designed， and the cDNA was used as a template for cloning the SE gene， which was analyzed for bioinformatics. The expression of P. cocos qualene epoxidase（PcSE） was examined by Real-time polymerase chain reaction（Real-time PCR） in P. coco Shenzhou No. 10， Xiangjing 28， and 5.78 strains. ResultThe full length of PcSE is 1 571 bp， containing four exons and three introns. The obtained CDS sequence is 1 413 bp， encoding 470 amino acids. This protein is a hydrophobic protein with no signal peptide structure and has two transmembrane structural domains with a FAD/NAD （P） binding domain and SE structural domain localized to the mitochondrial membrane and the plasma membrane. The homologous sequence alignment with fungi of the Poriferae family is 80.92%， and the phylogenetic tree shows that PcSE protein is most closely related to P. cocos from the US. The results of Real-time PCR showed that the PcSE was expressed in all three strains， with the highest expression in 5.78 strain， and there was no significant difference in PcSE expression among the three strains. ConclusionFor the first time， the PcSE gene was cloned and analyzed from P. cocos， providing a basis for further research on the function of PcSE and the analysis of P. cocos triterpene biosynthesis pathway.
