Search

Abstract: A reference karyotype is presented for loblolly pine (Pinus taeda L., subgenus Pinus, section Pinus, subsection Australes), based on fluorescent in situ hybridization (FISH), using 18S-28S rDNA, 5S rDNA, [...]

Physical mapping of BACs by fluorescent in situ hybridization (FISH) was used to analyze the liguleless (lg-1) linkage group in sorghum and compare it to the conserved region in rice and maize. Six liguleless-associated rice restriction fragment length polymorphism (RFLP) markers were used to select 16 homeologous sorghum BACs, which were in turn used to physically map the liguleless linkage group in sorghum. Results show a basic conservation of the liguleless region in sorghum relative to the linkage map of rice. One marker which is distal in rice is more medial in sorghum, and another marker which is found within the linkage group in rice is on a different chromosome in sorghum. BACs associated with linkage group I hybridize to chromosome [I.sub.t], which was identified by using FISH in a sorghum cytogenetic stock trisomic for chromosome I (denoted [I.sub.t]), and a BAC associated with linkage group E hybridized to an unidentified chromosome. Selected BACs, representing RFLP loci, were end-cloned for RFLP mapping, and the relative linkage order of these clones was in full agreement with the physical data. Similarities in locus order and the association of RFLP-selected BAC markers with two different chromosomes were found to exist between the linkage map of the liguleless region in maize and the physical map of the liguleless region in sorghum.

Genome-wide physical mapping with bacteria-based large-insert clones (e.g., BACs, PACs, and PBCs) promises to revolutionize genomics of large, complex genomes. To accelerate rice and other grass species genome research, we developed a genome-wide BAC-based map of the rice genome. The map consists of 298 BAC contigs and covers 419 Mb of the 430-Mb rice genome. Subsequent analysis indicated that the contigs constituting the map are accurate and reliable. Particularly important to proficiency were (1) a high-resolution, high-throughput DNA sequencing gel-based electrophoretic method for BAC fingerprinting, (2) the use of several complementary large-insert BAC libraries, and (3) computer-aided contig assembly. It has been demonstrated that the fingerprinting method is not significantly influenced by repeated sequences, genome size, and genome complexity. Use of several complementary libraries developed with different restriction enzymes minimized the 'gaps' in the physical map. In contrast to previous estimates, a clonal coverage of 6.0-8.0 genome equivalents seems to be sufficient for development of a genome-wide physical map of ~95% genome coverage. This study indicates that genome-wide BAC-based physical maps can be developed quickly and economically for a variety of plant and animal species by restriction fingerprint analysis via DNA sequencing gel-based electrophoresis.

Fluorescent in situ hybridization (FISH) of cloned probes provides a powerful tool for the integration of recombination, physical and cytogenetic maps. As part of a sorghum genomics project we have assigned the position of eight cloned DNA sequences to chromosome 1 of Sorghum bicolor. The probes used included a centromere-associated repeat, 28S-18S rDNA, a corn pollen-expressed Adh cDNA-selected sorghum bacterial artificial chromosome (BAC), and five RFLP-selected sorghum BACs. With the exception of the centromere-associated sequence and the 28S-18s rDNA, the probes produced FISH sites at the ends of the chromosome. The BACs selected from probes used to map two adjacent RFLP loci produced FISH signals at opposite ends of the chromosome. Over 85% of the physical length of this chromosome corresponded to approximately 12 map units separating the two RFLP loci. This indicates that most of the recombination and the loci of the ca. 120 cM RFLP map are located at the ends of the chromosome. Research supported by the Texas Advanced Technology and Research Program (grant 999902-090 to HJP and DMS), the Texas Agricultural Experiment Station, and the Texas A&M University Office of University Research.

Dvelops a genome-wide physical map of the rice genome based on the bacteria-based large-insert clones known as BAC. Map consisting of 298 BAC contigs covering 419 Mb; Electrophoretic method for BAC fingerprinting; Use of different restriction enzymes used in the physical map.