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Targeted expression of redesigned and codon optimised synthetic gene leads to recrystallisation inhibition and reduced electrolyte leakage in spring wheat at sub-zero temperatures

Khanna, Harjeet K. & Daggard, Grant (2006) Targeted expression of redesigned and codon optimised synthetic gene leads to recrystallisation inhibition and reduced electrolyte leakage in spring wheat at sub-zero temperatures. Plant Cell Reports, pp. 1336-1346.

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Abstract

Abstract Antifreeze proteins (AFPs) adsorb to ice crystals and inhibit their growth, leading to non-colligative freezing point depression. Crops like spring wheat, that are highly susceptible to frost damage, can potentially be made frost tolerant by expressing AFPs in the cytoplasm and apoplast where ice recrystallisation leads to cellular damage. The protein sequence for HPLC-6 α-helical antifreeze protein fromwinter flounderwas rationally redesigned after removing the prosequences in the native protein. Wheat nuclear gene preferred amino acid codons were used to synthesize a recombinant antifreeze gene, rAFPI. Antifreeze protein was targeted to the apoplast using a Murine leader peptide sequence from the mAb24 light chain or retained in the endoplasmic reticulum using C-terminus KDEL sequence. The coding sequences were placed downstream of the rice Actin promoter and Actin-1 intron and upstream of the nopaline synthase terminator in the plant expression vectors. Transgenic wheat lines were generated through micro projectile bombardment of immature embryos of spring wheat cultivar Seri 82. Levels of antifreeze protein in the transgenic lines without any targeting peptide were low (0.06–0.07%). The apoplast-targeted protein reached a level of 1.61% of total soluble protein, 90% of which was present in the apoplast. ER-retained protein accumulated in the cells at levels up to 0.65% of total soluble proteins. Transgenic wheat line T-8 with apoplast-targetedantifreeze protein exhibited the highest levels of antifreeze activity and provided significant freezing protection even at temperatures as low as −7◦C.

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10 citations in Web of Science®

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ID Code: 7276
Item Type: Journal Article
Keywords: Antifreeze protein, Codon optimisation, Frost tolerance, Signal peptide, Transgenic, Wheat
DOI: 10.1007/s00299-006-0191-9
ISSN: 1432-203X
Subjects: Australian and New Zealand Standard Research Classification > TECHNOLOGY (100000) > MEDICAL BIOTECHNOLOGY (100400) > Medical Molecular Engineering of Nucleic Acids and Proteins (100403)
Divisions: Past > QUT Faculties & Divisions > Faculty of Science and Technology
Current > Institutes > Institute of Health and Biomedical Innovation
Copyright Owner: Copyright 2006 Springer-Verlag
Copyright Statement: The original publication is available at SpringerLink http://www.springerlink.com
Deposited On: 02 May 2007
Last Modified: 29 Feb 2012 23:30

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