Abstract
Freshly isolated protoplasts from leaves of oat seedlings (var. Victory) which do not divide when cultured on a wide range of media are capable of incorporating tritiated leucine, uridine, and thymidine into trichloroacetic acid-insoluble macromolecules. Over 70% of the leucine and uridine incorporated over an 18-hour period are found in protein and RNA, respectively, as shown by hydrolysis of the macromolecular products with a specific protease or RNase. In contrast, little or none of the tritiated thymidine is incorporated into macromolecules hydrolyzable by DNase over an 18- to 96-hour period. Incorporation of thymidine into trichloroacetic acid-insoluble material declines sharply with increasing time of culture after 18 hours. However, addition of diamines or polyamines to the medium not only prevents the decline, but actually increases net thymidine incorporation, including a fraction going into DNA. A significant increase in mitoses and binucleate protoplasts is also observed in 72- to 168-hour cultures.
The inability of oat leaf protoplasts to synthesize significant quantities of DNA suggests that they are arrested at the G1 phase of the cell cycle. Treatment with polyamines appears to promote both DNA synthesis and the inception of mitotic activity in oat protoplasts, as in numerous animal and microbial cells.
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