Scientific Merit: The long term goal of this research project is to develop a single integrated cytological and genetic map of maize. This project will couple the vast cytological information available on the maize chromosomes with cutting edge 3D deconvolution fluorescence and confocal laser scanning microscopy. it is envisioned that this map will be a valuable resource to the maize community in particular and the grass community in general. Practical applications include a direct method to order of BACs and YACs. Basic questions that can be address with such a map include the determination of evolutionary relatedness between the grasses, the relationship between genetic and physical distance, the identification of recombination hot spots, the clustering of genes and repetitive DNA on chromosomes and the mapping of chromosomal domains that do not undergo recombination including the centromeres and the mapping of loci without discernible phenotypes. The proposed research is divided into three parts. The first will involve a determination of whether chromosomes condense uniformly during pachtene; the stage chosen as the basis for the cytological map because these offer a ten to fifteen fold increase in resolution over mitotic chromosomes and because they can be obtained in quantity. Condensation of chromosomes will be assessed throughout the five stages of pachytene by measuring the distances between cytological landmarks including heterochromatic knobs, nucleolar organizing regions, centromeres and telomeres. labelled probes are available for several of these landmarks. In the second part the knobs present in the parents of the maize Recombinant Inbred lines (RI lines) will be mapped cytologically using 3D FISH and genetically by scoring the knob composition in each of the 89 RI lines. In the third part, experiments are proposed to determine the cytological positions of genetically mapped genes by using either rice BACs as chromosome FISH probes to determine the position of several contiguous unique genes or by using single or double copy maize genes as FISH probes. If rice sequences prove too divergent, well characterized BACs from species more closely related to maize, such as sorghum, will be utilized.
