Identification of repetitive, genome-specific probes in crucifer oilseed species.
Direct amplification of minisatellite DNA by PCR (DAMD PCR) was used to amplify and subsequently clone several fragments of DNA from crucifer species. The PCR-derived fragments of DNA were generated using known minisatellite core sequences as PCR primers. Southern hybridization of these putative minisatellite DNA fragments revealed that many were genome-specific; they hybridized with high affinity only to the genomic DNA of the species from which they were cloned. The DNA fragments were believed to be dispersed in the genome, based on smear-like hybridization signals on EcoRI-, BamHI-, and HindIII-digested genomic DNA. Genome-specific probes were specifically isolated from Brassica rapa (A genome), Brassica nigra (B genome), and Sinapis alba in addition to several other crucifer species. The sequence of a B. rapa specific probe (pBr17.1.3A) contained a minisatellite region that could be divided into three tandem repeats; each repeat contained between two and five subrepeats and each subrepeat shared a highly conserved core region of 29 bp. This minisatellite sequence also hybridized with high affinity to the A genome species B. napus and B. juncea. This research showed that dispersed, genome-specific probes can be isolated using DAMD PCR and that these probes could be used to detect and quantify alien DNA present in progeny from intergeneric or interspecific crosses.
Somers
Daryl J
DJ
Agriculture and Agri-Food Canada, Cereal Research Centre, Winnipeg. SomersD@em.agr.ca
Demmon
Goewin
G
D001483:Base Sequence; D015139:Blotting, Southern; D019607:Brassicaceae / Q000235:genetics*; D003433:Crosses, Genetic; D015342:DNA Probes; D018744:DNA, Plant; D018745:Genome, Plant; D018598:Minisatellite Repeats; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction
D015342:DNA Probes; D018744:DNA, Plant