Preparation and sequencing of secreted proteins from the pharyngeal glands of the plant parasitic nematode Heterodera schachtii

Authors

  • Jan De Meutter,

    1. Vakgroep Moleculaire Genetica, Departement Plantengenetica, Vlaams Interuniversitair Instituut voor Biotechnologie (VIB), Universiteit Gent, K.L. Ledeganckstraat 35, B-9000 Gent, Belgium,
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  • Bartel Vanholme,

    1. Vakgroep Moleculaire Genetica, Departement Plantengenetica, Vlaams Interuniversitair Instituut voor Biotechnologie (VIB), Universiteit Gent, K.L. Ledeganckstraat 35, B-9000 Gent, Belgium,
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  • Gino Bauw,

    1. Vakgroep Moleculaire Genetica, Departement Plantengenetica, Vlaams Interuniversitair Instituut voor Biotechnologie (VIB), Universiteit Gent, K.L. Ledeganckstraat 35, B-9000 Gent, Belgium,
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  • Tom Tytgat,

    1. Instituut voor Dierkunde, Universiteit Gent, K.L. Ledeganckstraat 35, B-9000 Gent, Belgium
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  • Greetje Gheysen,

    1. Vakgroep Moleculaire Genetica, Departement Plantengenetica, Vlaams Interuniversitair Instituut voor Biotechnologie (VIB), Universiteit Gent, K.L. Ledeganckstraat 35, B-9000 Gent, Belgium,
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  • Godelieve Gheysen

    Corresponding author
    1. Vakgroep Moleculaire Genetica, Departement Plantengenetica, Vlaams Interuniversitair Instituut voor Biotechnologie (VIB), Universiteit Gent, K.L. Ledeganckstraat 35, B-9000 Gent, Belgium,
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    • a

      Present address: Vakgroep Moleculaire Biotechnologie, Faculteit Landbouwkundige en Toegepaste Biologische Wetenschappen, Universiteit Gent, Coupure links 653, B-9000 Gent, Belgium.


* Correspondence: Prof. G. Gheysen, Vakgroep Moleculaire Biotechnologie, Faculteit Landbouwkundige en Toegepaste Biologische Wetenschappen, Universiteit Gent, Coupure links 653, B-9000 Gent, Belgium. E-mail: godelieve.gheysen@rug.ac.be

summary

In order to gain insight into the biology of the parasitic relationship between Heterodera schachtii and its host plant, it is important to understand the functional role of the nematode’s pharyngeal secretions. These secretions presumably play a key role in establishing and maintaining a feeding site for the nematode. An optimized method was used for the in vitro production of H. schachtii pharyngeal gland secretions. These pharyngeal secretions were mainly produced in an insoluble form and could be solubilized under denaturing conditions for further analysis. The soluble fraction was concentrated with StrataClean (Stratagene, La Jolla, CA) or with a phenol/ether extraction. These methods made it possible for the first time to separate the secreted proteins on two-dimensional gels. By combining a micropreparative sample preparation with mass spectrometry, two β-1,4-endoglucanases were identified. A third spot was identified as a novel protein by microsequencing. This is the first report on protein sequence information from pharyngeal secretions of a plant parasitic nematode.

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