Abstract
The integration of genetic and physical maps of maize is progressing rapidly, but the cytogenetic maps lag behind, with the exception of the pachytene fluorescence in situ hybridization (FISH) maps of maize chromosome 9. We sought to produce integrated FISH maps of other maize chromosomes using Core Bin Marker loci. Because these 1 Kb restriction fragment length polymorphism (RFLP) probes are below the FISH detection limit, we used BACs from sorghum, a small-genome relative of maize, as surrogate clones for FISH mapping. We sequenced 151 maize RFLP probes and compared in silico BAC selection methods to that of library filter hybridization and found the latter to be the best. BAC library screening, clone verification, and single-clone selection criteria are presented along with an example of transgenomic BAC FISH mapping. This strategy has been used to facilitate the integration of RFLP and FISH maps in other large-genome species.
Highlights
Cytogenetics has proven invaluable in eukaryotic genome research, helping to elucidate genome structure in humans and model organisms such as fruit fly (Drosophila melanogaster) and maize (Zea mays) [1,2,3,4,5,6]
We determined the fulllength insert sequences (FLIS) of restriction fragment length polymorphism (RFLP) probes for markers that we selected for possible fluorescence in situ hybridization (FISH) mapping, including the Core Bin Markers (CBMs)
The maize RFLP-FLIS data turned out to be useful for maize genome annotation, we found that the use of the sequence information alone was not as productive as the conventional filter-hybridization method for identification of corresponding sorghum bacterial artificial chromosome (BAC) clones
Summary
Cytogenetics has proven invaluable in eukaryotic genome research, helping to elucidate genome structure in humans and model organisms such as fruit fly (Drosophila melanogaster) and maize (Zea mays) [1,2,3,4,5,6]. The field of cytogenetics, and in particular maize cytogenetics, was greatly advanced by Barbara McClintock’s pioneering work in the 1920s and 1930s [7]. Her method for unequivocal identification of individual chromosomes permitted major discoveries regarding the structure and dynamic behavior of the maize genome while establishing the connection between genetic and physical recombination [8,9,10]. In addition to singlelocus detection, fluorescence in situ hybridization (FISH) with repetitive sequence probes has been used and combined with multicolor cocktails to characterize genome structure in polyploid or closely related plant species [16,17,18,19]. Well-characterized libraries can serve as a resource for whole genome sequencing as was recently done for maize [21]
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