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Analysis of ELA-DQB Exon 2 Polymorphism in Argentine Creole Horses by PCR-RFLP and PCR-SSCP

Authors

  • E. E. Villegas-Castagnasso,

    1. Address of authors: Centro de Investigaciones en Genética Básica y Aplicada (CIGEBA), Facultad de Ciencias Veterinarias, Universidad Nacional de La Plata, 60 y 118 C.C. 296, C.P. B1900AVW, La Plata, Argentina;
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    • *

      These authors contributed equally to the present work. Fellows from CONICET.

  • S. Díaz,

    1. Address of authors: Centro de Investigaciones en Genética Básica y Aplicada (CIGEBA), Facultad de Ciencias Veterinarias, Universidad Nacional de La Plata, 60 y 118 C.C. 296, C.P. B1900AVW, La Plata, Argentina;
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  • G. Giovambattista,

    1. Address of authors: Centro de Investigaciones en Genética Básica y Aplicada (CIGEBA), Facultad de Ciencias Veterinarias, Universidad Nacional de La Plata, 60 y 118 C.C. 296, C.P. B1900AVW, La Plata, Argentina;
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  • F. N. Dulout,

    1. Address of authors: Centro de Investigaciones en Genética Básica y Aplicada (CIGEBA), Facultad de Ciencias Veterinarias, Universidad Nacional de La Plata, 60 y 118 C.C. 296, C.P. B1900AVW, La Plata, Argentina;
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  • P. Peral-García

    1. Corresponding author: Tel. fax: 54 221 4211799; E-mail: ppgarcia@fcv.unlp.edu.ar
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Centro de Investigaciones en Genética Básica y Aplicada (CIGEBA), Facultad de Ciencias Veterinarias, Universidad Nacional de La Plata, La Plata, Argentina

Summary

The second exon of equine leucocyte antigen (ELA)-DQB genes was amplified from genomic DNA of 32 Argentine Creole horses by PCR. Amplified DNA was analysed by PCR-restriction fragment length polymorphism (RFLP) and PCR-single-strand conformation polymorphism (SSCP). The PCR-RFLP analysis revealed two HaeIII patterns, four RsaI patterns, five MspI patterns and two HinfI patterns. EcoRI showed no variation in the analysed sample. Additional patterns that did not account for known exon 2 DNA sequences were observed, suggesting the existence of novel ELA-DQB alleles. PCR-SSCP analysis exhibited seven different band patterns, and the number of bands per animal ranged from four to nine. Both methods indicated that at least two DQB genes are present. The presence of more than two alleles in each animal showed that the primers employed in this work are not specific for a unique DQB locus. The improvement of this PCR-RFLP method should provide a simple and rapid technique for an accurate definition of ELA-DQB typing in horses.

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