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细胞色素P450是灵芝三萜生物合成途径中的关键元件,其催化反应过程中需要有NADPH/NADH为其传递电子。本课题组根据已获得的赤芝(Ganoderma lucidum(Leyss.Ex Fr.)Karst.)基因组数据中的CYP450和NADPH转录本序列,利用RT-PCR方法获得灵芝CYP450(GLCYP450)和NADPH(GLNADPH)基因的全长cDNA序列,并在序列两端引入相应的限制性内切酶位点,通过酶切及连接反应,将GLCYP450和GLNADPH重组到酵母表达载体pESC-URA中,成功构建了酵母表达载体质粒pESC-GLNADPH-GLCYP450,为通过合成生物学手段研究灵芝三萜生物合成奠定基础。
Cytochrome P450 is a key element in the biosynthesis pathway of ganoderma lucidum triterpenes, which needs NADPH / NADH to transmit electrons during its catalytic reaction. Based on the CYP450 and NADPH transcripts of Ganoderma lucidum (Leyss.Ex Fr.) Karst. Genome data obtained in the present study, we obtained the cDNAs of Ganoderma lucidum CYP450 (GLCYP450) and NADPH (GLNADPH) by RT-PCR The full-length cDNA sequence was introduced into the vector and introduced into the restriction enzyme site at both ends of the sequence. GLCYP450 and GLNADPH were recombined into the yeast expression vector pESC-URA by restriction enzyme digestion and ligation reaction. The yeast expression plasmid pESC -GLNADPH-GLCYP450, which laid the foundation for studying the biosynthesis of triterpenoids in Ganoderma lucidum by means of synthetic biology.