This chapter directs toward nucleolar chromosomes, their nucleolar organizers, and the origin of nucleolar substance. The role of the nucleolus in transmitting genetic information from the gene locus in the chromosome to the cytoplasm has been considered. The chapter also discusses the role of nucleolar chromosomes not only in normal cellular differentiation but also in the cancer process. In conventional cells, during interphase, only the nucleolar bodies are clearly visible as nuclear structures. In most cases, the chromosomes are invisible. Occasional allocycly provides a conspicuous chromatin mass that may be associated with the nucleolar bodies. The nucleolar chromosome consists of three integrated components: nucleolar body (nucleolus), the nucleolar organizer, and the chromosome. In some genomes, the nucleolar chromosome may carry considerable heterochromatin, especially for a sex chromosome. Experimental studies on nucleoli include modification of nucleoli by chemical agents, action of microbeam ultraviolet irradiation on nucleoli, modifications in nucleoli by genetics, and transplantation of nucleoli. The chapter gives a detailed account of lampbrush and polytene chromosomes. Experimental modifications of puffing patterns in salivary gland chromosomes have been provided in the chapter. The polytene chromosomes help to demonstrate functional activity of chromosomes and to obtain an insight into the mechanisms of transferring genetic information from chromosomes to cytoplasm. As these intriguing structures also have nucleoli, it helps to study the relationship between chromosome and nucleolus as well as between nucleolar and non-nucleolar chromosomes. The chapter also summarizes the relationships of chromosomes, nucleoli, nuclei, and other subcellular structures on the basic problems of cellular differentiation. With the availability of polytene chromosomes, coupled with experimental and genetic controls of puffing, the biologist has, for the first time, the material that pinpoints an operon going into action and controlling not only when the operon acts, but also of controlling the period of time the operon is permitted to act.