Cloning and prokaryotic expression analysis of DNA methyltransferas 1(MET1) in Ganoderma lucidum.
- Author:
Liang-Ping ZHA
1
;
Ya-Jun WANG
2
;
Shuang LIU
2
;
Yu-Yang ZHAO
2
;
Yuan YUAN
2
;
Lu-Qi HUANG
2
Author Information
- Publication Type:Journal Article
- Keywords: Ganoderma lucidum; gene cloning; methylation; methyltransferas; prokaryotic expression
- From: China Journal of Chinese Materia Medica 2016;41(6):1021-1026
- CountryChina
- Language:Chinese
- Abstract: DNA methyltransferase is the key enzyme in the process of DNA methylation, playing an important role in regulation of gene expression in vivo. According to the Ganoderma lucidum transcriptome data, a full-length cDNA sequence of MET1 from G. lucidum was cloned for the first time, the GenBank registration number is KU239998, and we conducted a comprehensive bioinformatics analysis of the genetic characteristics and spatial structure. The prokaryotic expression analysis showed that E.coli[pET28a(+)-GlMET1] in BL21(DE3) could induce objective protein, shaking the culture at 16 ℃ until the host bacterium(A₆₀₀) was approximately 0.8, and added IPTG to finally concentration of 0.2 mmol•L⁻¹, and then the optimal expression of GlMET1 recombinant protein was accumulated for the induction time of 20 h. The real-time PCR results showed that the expression levels of GlMET1 had obvious differences among varieties of G. lucidum. During the maturity stage, the expression levels of GlMET1 were lower than that in juvenile stage, the results showed that with the growth of G. lucidum, the expression levels of GlMET1 were on the decline. The research provided an important basis for studying the mechanism of DNA methyltransferase thoroughly.
