Scholarlink

In order to measure the actual synthesis and degradation rates (SR, DR) for rRNA in yeast, we developed a method based on the pulse labeling and quantification of newly synthesized large rRNA molecules by a known mass of cells. The SR is calculated as the ratio of new rRNA molecules (synthesized after a short [5,6-3H]-uracil pulse) to total rRNA (a proxy of cell mass), calculated by northern blotting after hybridization with a 32P-labeled rRNA probe. Then to measure the DR we perform a chase of the existing 3H-labeled rRNA for several hours during yeast culture growth. We have used this method in control experiments where the yeast cell volume varies as a way to check if the SR and DR are constant with the cell volume.

错误：搜索内容不能为空，请输入英文关键词
错误：关键词超出字数限制，请精简
高级检索

Measurement of rRNA Synthesis and Degradation Rates by 3H-Uracil Labeling in Yeast

  • 实验方案

  • 发表日期：

    2025年

查看全文
  • María J. García-Marcelo,
  • Gaurav Singh,
  • Sebastián Chávez,
  • José E. Pérez-Ortín

摘要

In order to measure the actual synthesis and degradation rates (SR, DR) for rRNA in yeast, we developed a method based on the pulse labeling and quantification of newly synthesized large rRNA molecules by a known mass of cells. The SR is calculated as the ratio of new rRNA molecules (synthesized after a short [5,6-3H]-uracil pulse) to total rRNA (a proxy of cell mass), calculated by northern blotting after hybridization with a 32P-labeled rRNA probe. Then to measure the DR we perform a chase of the existing 3H-labeled rRNA for several hours during yeast culture growth. We have used this method in control experiments where the yeast cell volume varies as a way to check if the SR and DR are constant with the cell volume.