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Cloning and expression analysis of candidate genes involved in wax deposition along the growing barley (Hordeum vulgare) leaf

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Abstract

The aim of the present study was to isolate clones of genes which are likely to be involved in wax deposition on barley leaves. Of particular interest were those genes which encode proteins that take part in the synthesis and further modification of very long chain fatty acids (VLCFAs), the precursors of waxes. Previously, it had been shown that wax deposition commences within a spatially well-defined developmental zone along the growing barley leaf (Richardson et al. in Planta 222:472–483, 2005). In the present study, a barley microarray approach was used to screen for candidate contig-sequences (www.barleybase.org) that are expressed particularly in those leaf zones where wax deposition occurs and which are expressed specifically within the epidermis, the site of wax synthesis. Candidate contigs were used to screen an established in-house cDNA library of barley. Six full-length coding sequences clones were isolated. Based on sequence homologies, three clones were related to Arabidopsis CER6/CUT1, and these clones were termed HvCUT1;1, HvCUT1;2 and HvCUT1;3. A fourth clone, which was related to Arabidopsis Fiddlehead (FDH), was termed HvFDH1;1. These clones are likely to be involved in synthesis of VLCFAs. A fifth and sixth clone were related to Arabidopsis CER1, and were termed HvCER1;1 and HvCER1;2. These clones are likely to be involved in the decarbonylation pathway of VLCFAs. Semi-quantitative RT-PCR confirmed microarray expression data. In addition, expression analyses at 10-mm resolution along the blade suggest that HvCUT1;1 (and possibly HvCUT1;2) and HvCER1;1 are involved in commencement of wax deposition during barley leaf epidermal cell development.

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Abbreviations

ACP:

Acyl-carrier protein

FAE:

Fatty acid elongase

FAS:

Fatty acid synthase

GL1:

Glossy 1

GL8:

Glossy 8

KCR:

3(β)-ketoacyl-reductase

KCS:

3(β)-Ketoacyl-CoA synthase

LACS:

Long-chain acyl-CoA synthetase

POE:

Point of emergence

POLI:

Point of leaf insertion

VLCFA:

Very long chain fatty acid

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Acknowledgements

This research was supported by the Biotechnology and Biological Sciences Research Council (BBSRC), UK, Grant 61/P18283, and benefited also from a research grant of The Leverhulme Trust, UK (to WF). We would like to thank Tobias Wojciechowski (who was funded through a studentship by University of Paisley) for his help with constructing the barley cDNA library and John Christie (Glasgow University) for use of the electroporator. We would also like to thank two anonymous referees for constructive comments on an earlier version of the manuscript.

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Author notes

  1. Andrew Richardson

    Present address: Millipore, Gemini Crescent, Dundee Technology Park, Dundee, DD2 1SW, UK

  2. Alexandre Boscari

    Present address: Interactions Plantes Microorganismes et Santé Végétale, UMR INRA 1064/Université de Nice-Sophia Antipolis/CNRS 6192, 400 Route des Chappes, B. P. 167, 06903, Sophia Antipolis Cedex, France

Authors and Affiliations

  1. Division of Biological Sciences, University of Paisley, Paisley, PA1 2BE, Scotland, UK

    Andrew Richardson, Alexandre Boscari & Wieland Fricke

  2. Department of Ecophysiology, IZMB, University of Bonn, 53115, Bonn, Germany

    Lukas Schreiber

  3. Department of Biological Sciences, Lancaster Environment Centre, Lancaster University, Lancaster, LA1 4YQ, UK

    Gerhard Kerstiens

  4. Department of Chemistry, University of Glasgow, Glasgow, G12 8QQ, Scotland, UK

    Mike Jarvis

  5. Sir Henry Wellcome Functional Genomics Facility, Institute of Biomedical and Life Sciences, University of Glasgow, Glasgow, G12 8QQ, UK

    Pawel Herzyk

  6. UCD School of Biology and Environmental Science, UCD Science Centre West, University College Dublin, Belfield, Dublin 4, Ireland

    Wieland Fricke

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Correspondence to Wieland Fricke.

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Richardson, A., Boscari, A., Schreiber, L. et al. Cloning and expression analysis of candidate genes involved in wax deposition along the growing barley (Hordeum vulgare) leaf. Planta 226, 1459–1473 (2007). https://doi.org/10.1007/s00425-007-0585-0

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