Study on mechanism of micronucleoli formation by laser microirradiation

In the male rat kangaroo cell line PTK2, argon laser (514.5 nm) microirradiation of both nucleoli in interphase cells 30, 23, and 12 h before mitosis, and nucleoli in early prophase cells resulted in the formation of micronucleoli, i.e., several small nucleolus-like bodies, in daughter cells. The irradiated cells were stained with methylene blue, which indicated that the nucleolar RNA was destroyed by laser microirradiation. Feulgen staining was applied to the irradiated cells in combination with the measurements of an MPV-II model microphotometer. Irradiated nucleoli were negative for DNA-Feulgen stain, which indicated that nucleolar DNA was destroyed by laser irradiation, so the nucleolar organizer gene was destroyed. After the nucleoli had been irradiated, the cells were continuously incubated at 37 degrees C for 12 and/or 24 h, then fixed and stained with AgNO3. Most of the nucleoli irradiated silver-stained negative that demonstrated that when the nucleoli were irradiated, rDNA was destroyed and transcription stopped. However, some silver grains were found in the nucleoplasm, whereas the nucleoli were silver-stained negative. The results suggest that subsidiary nucleolar organizer loci might exist scattered throughout the genome.