Browsing by Subject "BAC library"
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Item Comparative Mapping of the Alpaca Genome(2014-08-06) Fagundes De Avila, FelipeThe development of gene maps constitutes a key feature for understanding genome architecture and comparative evolution. The genomes of some livestock species such as cattle, horses and pigs, have received considerable attention over the years due to their economic importance. In contrast, though camelids are gaining worldwide popularity as production and companion animals, cytogenetics and genome mapping in these species lag far behind those of other mammals. One of the reasons for the scarce body of knowledge regarding the camelid genome is their particularly difficult karyotype for analysis. All six extant camelid species have a diploid number of 74 chromosomes; the gross morphological similarities shared by many of the autosomes, combined with the relatively small size of some chromosome pairs, present serious challenges for identifying individual chromosomes using conventional cytogenetic techniques. The Alpaca Genome Project includes whole genome sequencing, radiation hybrid (RH) mapping and human-camel comparative chromosome painting (Zoo-FISH). However, there is no common platform that aligns various maps and precisely assigns them to individual chromosomes. Therefore, the goal of this research project was to construct a cytogenetic map for the alpaca genome by fluorescence in situ hybridization (FISH) of large insert clones from the alpaca CHORI-246 genomic BAC library. The BACs were selected based on the available Zoo-FISH, RH and sequence map data to target evolutionarily conserved genes and to get uniform distribution of markers throughout the alpaca genome. Candidate genes for traits of interest such as various congenital and reproduction-related disorders, as well as for phenotypic traits such as fiber color and texture, were also selected for mapping. A total of 230 markers were mapped to the 36 alpaca autosomes and the sex chromosomes; moreover, comparative mapping showed exceptional conservation of both gene synteny and order between alpaca and dromedary camel chromosomes. The cytogenetic map of the alpaca genome is a platform that effectively integrates the whole genome sequence and the radiation hybrid map with cytogenetic data, thus facilitating the discovery of genes of interest and providing tools for studying chromosome evolution and for clinical cytogenetics by means of a collection of chromosome-specific markers for camelids.Item Genomic organization of chromosomal centromeres in the cultivated rice, Oryza sativa L., and its wild progenitor, O. rufipogon Griff.(Texas A&M University, 2004-11-15) Uhm, TaesikCentromeres are responsible for sister-chromatid cohesion, kinetochore formation, and accurate transmission of chromosomes. Rice provides an excellent model for organizational and functional studies of centromeres since several of its chromosomes contain limited amounts of satellite and other repetitive sequences in their centromeres. To facilitate molecular characterization of the centromeres, we screened several BIBAC and BAC libraries of japonica and indica rice, using several centromere-specific repeat elements as probes. The positive clones were identified, fingerprinted and integrated into our whole genome physical map databases of the two rice subspecies. BAC/BIBACbased physical maps were constructed for the centromeric regions of the subspecies. To determine whether the genomic organization of the centromeres has changed since the cultivated rice split from its progenitor and to identify the sequences potentially playing an important role in centromere functions, we constructed a large-insert BIBAC library for the wild progenitor of Asian cultivated rice, O. rufipogon. The library contains 24,192 clones, has an average insert size of 163 kb, and covers 5 x haploid genome of wild rice. We screened the wild rice library with two centromere 8-specific overgo probes designed from the sequences flanking centromere 8 of japonica rice. A BIBACbased map was constructed for wild rice centromere 8. Two of the clones, B43P04 and B15E04, were found to span the entire region of the wild rice centromere and thus selected for sequencing the centromere. By sequencing the B43P09 clone, a 95% genomic sequence of the long arm side of wild rice centromere 8 was obtained. Comparative analysis revealed that the centromeric regions of wild rice have a similar gene content to japonica rice, but the centromeric regions of japonica rice have undergone chromosomal rearrangements at both large scale and nucleotide levels. In addition, although the 155-bp satellite repeats showed dramatic changes at the middle region, they are conserved at the 5' and 3' ends of satellite monomers, suggesting that those regions might have important functional roles for centromeres. These results provide not only new insights into genomic organization and evolution, but also a platform for functional analysis of plant centromeres.