Myoglobin intron variation in the Gouldian Finch Erythrura gouldiae assessed by temperature gradient gel electrophoresis

Authors

  • Margaret M. Heslewood,

    Corresponding author
    1. Centre for Conservation Technology, School of Resource Science and Management, Southern Cross University, Lismore, NSW, Australia
    • Centre for Conservation Technology, School of Resource Science and Management, Southern Cross University, P.O. Box 157, Lismore NSW 2480, Australia (Tel: +61-2-662-03815; Fax: +61-2-662-12669)
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  • Martin S. Elphinstone,

    1. Centre for Conservation Technology, School of Resource Science and Management, Southern Cross University, Lismore, NSW, Australia
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  • Sonia C. Tidemann,

    1. Conservation Commission of the Northern Territory, Palmerston, NT, Australia
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  • Peter R. Baverstock

    1. Centre for Conservation Technology, School of Resource Science and Management, Southern Cross University, Lismore, NSW, Australia
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Abstract

In this study we show how the use of exon-primed, intron-crossing (EPIC) polymerase chain reaction (PCR) of a diploid intronic region, in conjunction with temperature gradient gel electrophoresis (TGGE), can be used to detect and rapidly assess allelic variation at the nucleotide level. We developed passerine-specific primers to amplify and sequence a 762 bp region including the second intron of the myoglobin gene in the Gouldian Finch, Erythrura gouldiae. A POLAND plot based on this sequence indicated that TGGE in combination with heteroduplex analysis (TGGE/HA) should reveal nucleotide variation in the 160 bp low-melting domain. Sequencing of the entire fragment from 19 Er. gouldiae revealed five nucleotide substitution differences within the low-melt domain, all of which could be detected and differentiated by TGGE/HA, and an additional substitution in a section of the high-melt domain which characterised another allele. A total of 181 individuals from four populations were screened for these six alleles.

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