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Molecular Cloning, Characterisation, and Heterologous Expression of Farnesyl Diphosphate Synthase from Sanghuangporus baumii

Molecular Biotechnology volume 62, pages 132–141 (2020)Cite this article

Abstract

A farnesyl diphosphate synthase (FPS) cDNA and promoter region was cloned from Sanghuangporus baumii. The gene contains a 150-bp 5′-untranslated region (UTR), a 154-bp 3′-UTR, and a 1062-bp open reading frame (ORF) encoding a 354 amino acid polypeptide. The FPS-DNA includes three exons (nucleotides 1 –123, 184–321, and 505–1305) and two introns (nucleotides 124–183 and 322–504). The FPS protein has a molecular weight of 40.73 kDa, it is hydrophilic with a theoretical isoelectric point of 5.13, and the secondary and three-dimensional structure were analysed. There is a transcription start site at nucleotides 1318–1368 of the promoter, which includes typical eukaryotic promoter elements (TATA Box, CAAT Box, ARBE, AT-rich element, G-box, MBS, Sp1, LTR). FPS was expressed in Escherichia coli BL21, and the recombinant protein (63.41 kDa) was subjected to dodecyl sulphate, sodium salt-polyacrylamide gel electrophoresis (SDS-PAGE). FPS transcription was measured during different developmental stages, and expression in 11 and 13 days mycelia was upregulated 49.3-fold and 125.4-fold, respectively, compared with 9 days mycelia controls. Through analysing, S. baumii triterpenoid content was correlated with the transcription level of FPS during different development stages, and the triterpenoid content peaked at day 15 (7.21 mg/g).

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Acknowledgements

This research was supported by the Fundamental Research Funds for the Central Universities, China (2572017CF01).

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Affiliations

  1. College of Forestry, Northeast Forestry University, Hexing Road 26, Xiangfang District, Harbin, 150040, Heilongjiang, China

    Xutong Wang, Jian Sun, Shixin Wang, Yisha Ma, Zengcai Liu, Jian Zhang, Guoquan Zhang & Li Zou

  2. Department of Food Engineering, Harbin University, Zhongxing Road 109, Nangang District, Harbin, 150086, Heilongjiang, China

    Tingting Sun

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Wang, X., Sun, T., Sun, J. et al. Molecular Cloning, Characterisation, and Heterologous Expression of Farnesyl Diphosphate Synthase from Sanghuangporus baumii. Mol Biotechnol 62, 132–141 (2020). https://doi.org/10.1007/s12033-019-00231-0

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Keywords

  • Sanghuangporus baumii
  • Farnesyl diphosphate synthase
  • Triterpenoids
  • Promoter
  • Intron analysis
  • Quantitative real-time PCR
  • Recombinant expression
  • Bioinformatics