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Molecular Cloning, Bacterial Overexpression and Characterization of L-myo- inositol 1- Phosphate Synthase from a Monocotyledonous Resurrection Plant, Xerophyta viscosa Baker

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Abstract

L-myo-inositol 1-phosphate synthase (EC 5.5.1.4; MIPS), an evolutionarily conserved enzyme-protein, catalyses the first and rate limiting step of inositol biosynthesis. Inositol and its derivatives play important roles in biological kingdom like growth regulation, membrane biogenesis, signal transduction and also acts as an osmolyte or osmoprotectant in abiotic stress tolerance. Here we report the cloning, sequencing and the characterization of the INO1 gene from Xerophyta viscosa (XINO1), a monocotyledonous resurrection plant. Nucleotide sequences of XINO1 show striking homology (70–99%) with a number of INO1 genes from plant sources particularly with the monocots. The gene is functionally identified through genetic complementation using a yeast inositol auxotrophic strain FY250. The gene is expressed in E. coli BL21, recombinant protein purified to homogeneity, biochemically characterized and compared with Oryza INO1 (RINO1) gene product. The XINO1 gene product is catalytically active in a broader range of lower temperature (between 10–40 °C) than the RINO1 gene- product. This is the first report of MIPS gene from any resurrection plant.

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Abbreviations

IPTG:

Isopropyl β-D-thiogalactopyranoside

PMSF:

Phenylmethylsulfonyl fluoride

G 6 P:

D-Glucose 6-Phosphate

X-gal:

5-Bromo-4-Chloro-3Indolyl-Beta-D-Thiogalactopyranoside

β-ME:

β-Mercaptoethanol

NAD:

Nicotinamide adeninedinucleotide

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Author notes

  1. Manoj Majee

    Present address: Department of Horticulture, University of Kentucky, Lexington, USA

Authors and Affiliations

  1. Plant Molecular and Cellular Genetics, Bose institute (Centenary Building), P-1/12, CIT Scheme VIIM, Kolkata, 700 054, India

    Manoj Majee, Barunava Patra & Arun Lahiri Majumder

  2. University of Cape Town, Cape Town, South Africa

    Sagadevan G. Mundree

Authors
  1. Manoj Majee
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  2. Barunava Patra
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  3. Sagadevan G. Mundree
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  4. Arun Lahiri Majumder
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Corresponding author

Correspondence to Arun Lahiri Majumder.

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Majee, M., Patra, B., Mundree, S.G. et al. Molecular Cloning, Bacterial Overexpression and Characterization of L-myo- inositol 1- Phosphate Synthase from a Monocotyledonous Resurrection Plant, Xerophyta viscosa Baker. J. Plant Biochem. Biotechnol. 14, 95–99 (2005). https://doi.org/10.1007/BF03263235

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