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Cloning and heterologous expression of isopentenyl diphosphate isomerase gene from Lycium chinense

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Abstract

Isopentenyl diphosphate isomerase (IPI), a key enzyme of terpenoid biosynthesis pathway, catalyzes the reversible conversion between isopentenyl diphosphate (IPP) and its allylic isomer dimethylallyl diphosphate (DMAPP), which are essential precursors for terpenoid biosynthesis. In this study, we isolated and characterized the IPI gene from Lycium chinense (LcIPI). The full-length cDNA sequence was 1104 bp encoding 293 amino acid residues. Additionally, several specific conserved motifs of IPI were also identified by amino acid sequence analysis in this protein. Phylogenetic tree showed that LcIPI had a close relationship with other solanaceae plants. Expression of recombinant protein in Escherichia coli produced a clear target band of about 35 kDa. Enzymatic activity was determined by heterologous complementation analysis, and β-carotene content of LcIPI transformants was about two times more compared with control. Furthermore, quantitative real-time PCR assay demonstrated that LcIPI was expressed in all tissues tested with the highest level in flowers. The results obtained here clearly suggest that the LcIPI gene is a promising candidate for carotenoid production in heterologous hosts.

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Abbreviations

IPP:

Isopentenyl diphosphate

DMAPP:

Dimethylallyl diphosphate

IPI:

Isopentenyl diphosphate isomerase

HPLC:

High-performance liquid chromatography

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Acknowledgments

The authors appreciate the kind presentation of plasmid pACCAR16ΔcrtX from professor Chanfu Zhu (School of Life Sciences, Northeast Normal University, Changchun, China). This work was financially supported by the National Science and Technology Key Project of China on GMO Cultivation for New Varieties (No. 2014ZX08003-002B).

Conflict of interest

The authors declare that they do not have any conflict of interest.

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Authors and Affiliations

  1. School of Chemical Engineering and Technology, Tianjin University, Tianjin, 300072, People’s Republic of China

    Zhaodi Li, Tchouopou Lontchi Josine, Jiang Wu, Jinjin Diao & Weidang Wu

  2. School of Environmental Science and Engineering, Tianjin University, Tianjin, 300072, People’s Republic of China

    Jing Ji, Gang Wang & Chunfeng Guan

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  1. Zhaodi Li
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  2. Jing Ji
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  3. Gang Wang
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  4. Tchouopou Lontchi Josine
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  5. Jiang Wu
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  6. Jinjin Diao
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  7. Weidang Wu
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  8. Chunfeng Guan
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Corresponding author

Correspondence to Jing Ji.

Electronic supplementary material

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Supplementary 1

Partial sequence of isopentenyl diphosphate isomerase gene from Lycium chinense. (DOC 651 kb)

Supplementary 2

Three-dimensional model of the Lycium chinense IPI. The ribbons and arrows stand for α-helix and β-sheet regions, respectively. The loop regions are also present in the schematics. (DOC 4108 kb)

Supplementary 3

Primers sequence of Arabidopsis thaliana and Zea mays. Underline stands for the restriction site of BamHI and XholI respectively. (DOC 37 kb)

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Li, Z., Ji, J., Wang, G. et al. Cloning and heterologous expression of isopentenyl diphosphate isomerase gene from Lycium chinense . J. Plant Biochem. Biotechnol. 25, 40–48 (2016). https://doi.org/10.1007/s13562-015-0306-x

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  • DOI: https://doi.org/10.1007/s13562-015-0306-x

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