Abstract
To examine gene expression, Northern blot or Real-Time PCR can be used to detect low abundant RNA such as mRNA. However, for high abundant RNAs such as rRNA and tRNA, Northern blot will not be able to discriminate the newly synthesized RNA from total RNA. Therefore, metabolic labeling is necessary to evaluate the expression of rRNA and tRNA genes. In this protocol, I describe a step-by-step method for labeling yeast RNA with radioactive uracil and examine the synthesis of these high abundant RNAs.
Keywords: Autoradiography, Metabolic labeling, Tritium, RRNA, TRNA
Materials and Reagents
Equipment
Procedure
Recipes
Acknowledgments
This protocol is derived from the following papers, Wei et al. (2009a) and Wei et al. (2009b), and the relevant references therein. The work was supported by NIH grants R01-CA099004 and R01-CA123391 to Dr. X.F. Steven Zheng at Robert Wood Johnson Medical School, University of Medicine and Dentistry of New Jersey.
References
If you have any questions/comments about this protocol, you are highly recommended to post here. We will invite the authors of this protocol as well as some of its users to address your questions/comments. To make it easier for them to help you, you are encouraged to post your data including images for the troubleshooting.