Hsp90 gene, an additional target for discrimination between the potato cyst nematodes, Globodera rostochiensis and G. pallida, and the related species, G. tabacum tabacum |
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Authors: | Mehrdad Madani Len J Ward and Solke H De Boer |
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Institution: | (1) Charlottetown Laboratory, Canadian Food Inspection Agency, 93 Mount Edward Road, C1A 5T1 Charlottetown, PE, Canada |
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Abstract: | The heat-shock gene, Hsp90, was targeted as a new variable genomic region to supplement other DNA-based tests for identification and discrimination
of Globodera pallida, G. rostochiensis and G. tabacum tabacum. Populations of the potato cyst nematodes, G. pallida and G. rostochiensis (PCN), originating from Canada, France, Belgium and USA, together with two populations of G. tabacum tabacum from the USA and France were used for the amplification of a fragment of the Hsp90 gene. General and specific primers and probes for each species were derived from the consensus and non-consensus regions
of the aligned sequences, respectively. A triplex conventional PCR assay, using a general forward and reverse or three specific
reverse primers, as well as a real-time PCR using general primers and specific TaqMan probes, were developed. Melting curve
analysis and restriction fragment polymorphisms using high resolution electrophoresis were explored for identifying PCR amplicons
that characterized and discriminated the three Globodera species in both pure and mixed samples. Results from the different molecular assay strategies confirmed the usefulness of
Hsp90 as a new additional gene target and showed that several different test options could be used for discrimination of PCN. |
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