The goal of the research is to develop a method to generate ordered sets of chromosome deletions. Plant chromosome arms will be translocatcd into the genome of cultured cell lines via Cre-lox site-specific recombination. The resulting hybrid chromosomes will be referred to as RACs. Subclone cell lines will be generated carrying deletion derivatives of the plant chromosome arm. The deletions will be made through homology-dependent gene targeting of new lox sites downstream of the translocation breakpoint, followed by the deletion of intervening DNA between the two lox sites. An ordered set of deletions for each plant chromosome arm can be generated with the insertion of a series of downstream lox sites. For a RAC deletion library of the maize genome, the goal will be to insert new lox sites at ~2.5 x 107 bp apart, resulting in ~200 RACs, each containing a maize chromosome fragment of 25 to 250 Mbp with defined endpoints on the maize chromosome map. The RAC deletion library can be used for physical mapping of Yeast or Bacterial Artificial Chromosome (YAC or BAC), or individual cDNAs. The specific objectives of the current project are first, to develop the method of generating RACs in cultured cells, and second, to generate maize plants with a lox site on each chromosome arm near the centromere.
Results from this research will provide a valuable tool for the physical mapping of the maize genome and other large genomes. Such a tool will allow detailed analysis of large plant genomes in a way that has not been possible before.
