The cellular slime mold Dictyostelium discoideum is a lower eukaryotic social amoeba with a simple developmental life cycle. The genes coding for ribosomal RNA in Dictyostelium are present in 200 copies per haploid genome, and are located on a giant linear extrachromosomal DNA molecule (rDNA) which is 80,000 base pair long. The DNA sequence of each extrachromosomal copy of the rDNA appears to be a perfect palindrome: two identical 40,000 base pair sequences joined tail to tail to form a linear inverted repeat. Two 6,000 base pair ribosomal RNA transcription units are symmetrically located near either end of the rDNA molecule, and are separated by 60,000 base pairs of "spacer" DNA of unknown function. We propose to study the structure of the extrachromosomal rDNA in Dictyostelium at the DNA sequence level. Through a combination of restriction enzyme mapping, electron microscopy, nucleic acid hybridization, RNA and DNA sequence determination, and molecular cloning using recombinant DNA techniques, we can approach the following questions: 1. How do these linear extrachromosomal elements replicate? 2. Are the extrachromosomal rDNA molecules derived from one or more integrated chromosomal copies by a process of gene amplification at some stage in the Dictyostelium life cycle? 3. Does the 60,000 base pair spacer sequence code for any structural or messenger RNA species? 4. Are any non-coding or even coding sequences within the extrachromosmal rDNA homologous to integrated chromosomal loci elsewhere in the genome, as we might expect for transcriptional control elements shared by many genes, or for sequences essential for initiation and termination of DNA replication? 5. Will a detailed structural analysis of the rDNA reveal unusual features such as interruptor or intervening sequences, internal palindromes within the larger palindrome, heterogeneity in either length or sequence, or pseudogenes, as have been found in the rDNA of other organisms?