Title: The applicability of consensus PCR primers across species and genera: the use of wheat Em sequences to develop markers for orthologues in rye
Authors: Van Campenhout, Steven ×
Koebner, RMD
Volckaert, Guido #
Issue Date: 2000
Publisher: Springer verlag
Series Title: Theoretical and applied genetics vol:100 issue:2 pages:328-336
Abstract: Two wheat consensus primer sets, directed to "early-methionine-labelled" (Em) gene sequences, were tested for their ability to amplify beyond their original source. A range of widely diverse templates, including other Triticeae species and sample monocot and dicot species, was assayed. Primer set EMC5/EMC3, amplifying the entire coding region with its intron and part of the 3' untranslated region, targets Triticeae and sorghum Fm sequences. The other set, EMC5/EMC031, directed to the coding region and its intron, amplifies templates from all the grass species. Both primer sets fail to amplify Fm sequences from more distant monocots and the dicots. Using set EMC5/EMC3, we isolated and sequenced ten members of the rye Em gene family from five differ ent rye sources. Significant DNA sequence variation between wheat and rye sequences in the non-coding regions was found, and this was used to develop seven sequence-specific primers. Twelve primer combinations were analysed, 7 of which were Em-R1-specific, amplifying a product in at least one of the tested rye or rye-carrying genotypes but not in wheat. Four sets exhibited clear amplification length polymorphisms which allowed discrimination between and within the Die sources. The primers also discriminated between wheat-rye recombinants with proximal 1RL rye chromatin and those carrying distal 1RL rye chromatin. These results show that wheat consensus primer sets can be used to isolate orthologous sequences, especially from species that are used for alien gene transfer in wheat. Subsequently, species-specific assays can be designed that are useful tools for this application.
ISSN: 0040-5752
Publication status: published
KU Leuven publication type: IT
Appears in Collections:Division of Gene Technology (-)
× corresponding author
# (joint) last author

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