Cloning and characterization of a squalene synthase gene from a medicinal mushroom, Inonotus obliquus(Fr.) Pilat
会议名称：《中国生物工程学会2014年学术年会暨全国生物技术大会》
会议日期：2014年
学科分类：1008[医学-中药学(可授医学、理学学位)] 0710[理学-生物学] 07[理学] 071007[理学-遗传学] 10[医学]
关 键 词：Squalene synthase gene Inonotus obliquus Medicinal mushroom Promoter Methyl jasmonate Total triterpenoids
摘 要：Squalene synthase catalyzes the condensation of two molecules of farnesyl diphosphate to produce squalene, the first committed precusor for sterol, brassinosteroid, and triterpene biosynthesis. A squalene synthase gene, designated Io SQS, was isolated from Inonotus obliquus, a medicinal mushroom produces a plethora of bioactive triterpenes. The Io SQS cDNA was found to contain an open reading frame of 1,476 bp, encoding a protein of 491 amino acids with a calculated molecular mass of 55.85 kDa. The Io SQS genomic DNA sequence consisted of 1,813 bp and contained four exons and three introns. The restriction fragment polymorphisms revealed by Southern blot analysis suggested that IoSQS was a single copy gene. Promoter analysis indicated that the 5’ upstream region of IoSQS possessed various potential elements associated with physiological and environmental factors. The expression pattern of IoSQS in different stages and under MeJA treatment were correlative with the accumulation of total triterpenoids, and was consistent with the predicted results of Io SQS promoter region. The N-terminal 466 residues of the hydrophilic sequence was expressed as a His-tagged protein in E. coli, and the resultant bacterial crude extract was incubated with farnesyl diphosphate and NADPH, squalene was detected in vitro reaction mixture by HPLC analysis. These results suggest that IoSQS enzyme is involved in squalene production in I. obliquus.