title: Effective isolation of retrotransposons and repetitive DNA families from the wheat genome.
abstract: New classes of repetitive DNA elements were effectively identified by isolating small fragments of the elements from the wheat genome. A wheat A genome library was constructed from Triticum monococcum by degenerate cleavage with EcoO109I, the recognition sites of which consisted of 5'-PuGGNCCPy-3' multi-sequences. Three novel repetitive sequences pTm6, pTm69 and pTm58 derived from the A genome were screened and tested for high copy number using a blotting approach. pTm6 showed identity with integrase domains of the barley Ty1-Copia-retrotransposon BARE-1 and pTm58 showed similarity to the barley Ty3-gypsy-like retrotransposon Romani. pTm69, however, constituted a tandem array with useful genomic specificities, but did not share any identity with known repetitive elements. This study also sought to isolate wheat D-genome-specific repetitive elements regardless of the level of methylation, by genomic subtraction. Total genomic DNA of Aegilops tauschii was cleaved into short fragments with a methylation-insensitive 4 bp cutter, MboI, and then common DNA sequences between Ae. tauschii and Triticum turgidum were subtracted by annealing with excess T. turgidum genomic DNA. The D genome repetitive sequence pAt1 was isolated and used to identify an additional novel repetitive sequence family from wheat bacterial artificial chromosomes with a size range of 1 395-1 850 bp. The methods successfully led pathfinding of two unique repetitive families.
authors.affiliation: Molecular Genetics Laboratory, Faculty of Agriculture, Tottori University, Tottori 680-8553, Japan. tomita206@gmail.com
authors.forename: Motonori
authors.initials: M
authors.lastname: Tomita
authors.forename: Munenori
authors.initials: M
authors.lastname: Asao
authors.forename: Aya
authors.initials: A
authors.lastname: Kuraki
mesh_terms: D015139:Blotting, Southern; D018744:DNA, Plant / Q000235:genetics*; D018745:Genome, Plant / Q000235:genetics*; D010802:Phylogeny; D010940:Plant Proteins / Q000145:classification / Q000235:genetics; D012091:Repetitive Sequences, Nucleic Acid / Q000235:genetics*; D018626:Retroelements / Q000235:genetics*; D017422:Sequence Analysis, DNA; D014908:Triticum / Q000235:genetics*
chemical_list: D018744:DNA, Plant; D010940:Plant Proteins; D018626:Retroelements