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

Khanna, Harjeet K. and Daggard, Grant E. (2006) Targeted expression of redesigned and codon optimised synthetic gene leads to recyrstallisation inhibition and reduced electrolyte leakage in spring wheat at sub-zero temperatures. Plant Cell Reports, 25 (12). pp. 1336-1346. ISSN 0721-7714

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-targeted antifreeze 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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Item Type: Article (Commonwealth Reporting Category C)
Refereed: Yes
Item Status: Live Archive
Additional Information: Awaiting Author's version. Deposited in accordance with the copyright policy of the publisher.
Depositing User: epEditor USQ
Faculty / Department / School: Historic - Faculty of Sciences - No Department
Date Deposited: 16 Nov 2007 05:30
Last Modified: 02 Jul 2013 22:54
Uncontrolled Keywords: antifreeze protein, codon optimisation, frost tolerance, signal peptide, transgenic, wheat
Fields of Research (FOR2008): 10 Technology > 1001 Agricultural Biotechnology > 100105 Genetically Modified Field Crops and Pasture
07 Agricultural and Veterinary Sciences > 0703 Crop and Pasture Production > 070305 Crop and Pasture Improvement (Selection and Breeding)
Identification Number or DOI: doi: 10.1007/s00299-006-0191-9
URI: http://eprints.usq.edu.au/id/eprint/3541

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