Guo Xu, Sun Chao, Song Jing-Yuan, Luo Hong-Mei, Chen Shi-Lin
National Engineering Laboratory for Breeding of Endangered Medicinal Materials, Institute of Medicinal Plant Development, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing 100193, China.
Yao Xue Xue Bao. 2013 Feb;48(2):206-10.
Cytochrome P450 (CYP450) is a key element in the Ganoderma triterpenoid biosynthetic pathway. The catalytic reaction process for CYP450 requires NADPH / NADH for electron transfer. After searching the genome dataset of Ganoderma lucidum, the unique sequence encoding CYP450 and NADPH were discovered, separately. The open reading frames of GLCYP450 and GLNADPH were cloned separately using RT-PCR strategy from G lucidum. The appropriate restriction enzyme cutting sites were introduced at the 5' and 3' ends of gene sequence. The genes of GLCYP450 and GLNADPH were recombined into the yeast expression vector pESC-URA, leading to the formation of the yeast expression plasmid pESC-GLNADPH-GLCYP450. This study provides a foundation for researching Ganoderma triterpene biosynthesis using the approach of synthetic biology.
细胞色素P450（CYP450）是灵芝三萜生物合成途径中的关键要素。CYP450的催化反应过程需要NADPH / NADH进行电子转移。在搜索灵芝基因组数据集后，分别发现了编码CYP450和NADPH的独特序列。采用RT-PCR策略从灵芝中分别克隆了GLCYP450和GLNADPH的开放阅读框。在基因序列的5'和3'末端引入了合适的限制性内切酶切割位点。将GLCYP450和GLNADPH基因重组到酵母表达载体pESC-URA中，从而形成酵母表达质粒pESC-GLNADPH-GLCYP450。本研究为利用合成生物学方法研究灵芝三萜生物合成提供了基础。
