Browsing by Subject "FISH"
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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 analysis of sorghum by fluorescence in situ hybridization(Texas A&M University, 2004-11-15) Kim, Jeong-SoonThe reliability of genome analysis and proficiency of genetic manipulation in vivo and in vitro are increased by assignment of linkage groups to specific chromosomes, placement of centromeres, orientation with respect to telomeres, and linear alignment with respect to chromosomal features and dimensions. I undertook five studies aimed at integrating sorghum genomics and cytogenetics at several levels. The results help establish an entirely new "cyto-genomics" resource, impacts of which are likely to be broad. In the first study, I developed a FISH-based karyotyping system for Sorghum bicolor Moench. I used integrated structural genomic resources, including linkage maps and large-insert clonal libraries of sorghum genomic DNA to develop a 17-locus probe cocktail for simultaneous fluorescent in situ hybridization (FISH). This probe enabled facile identification of all chromosome pairs in mitotic chromosome spreads. Perhaps just as important, I established time-efficient means to select sorghum BAC clones for multi-probe FISH. Thus, an integrated cyto-genomics system for sorghum can be constructed without need of chromosome flow sorting or microdissection, both of which are difficult and costly. In the second study, hybridization of DNA clones from 37 different genomic regions enabled the assignment of linkage groups and orientation of linkage maps to chromosomes. Comparisons between genetic and physical distances throughout the genome enabled a new nomenclature for linkage group designation in sorghum. The results provide an integrated nomenclature system of Sorghum bicolor chromosomes and linkage groups. In the third study, I created high-resolution maps by FISH to pachytene bivalents for two linkage groups (B and H), and defined relationships between pericentromeric heterochromatin, centromeres, mapped markers and recombination rates. These relationships will help guide the development and use of sorghum genomics. In the fifth study, I used FISH in two ongoing gene-targeted efforts. For the maturity gene ma5 and fertility restoration gene rfl, I estimated physical lengths between currently available flanking molecular markers. This enables estimation of recombination densities in these regions and assessment of the applicability of map-based and -assisted cloning.