A technique is described to directly measure ongoing transcription from 
individual genes in permeabilized cells of either the budding yeast 
Saccharomyces cerevisiae or the fission yeast Schizosaccharomyces pombe. 
Transcription run-on (TRO) analysis is used to compare the relative rates of 
synthesis for specific transcripts in cells grown under different environmental 
conditions or harvested at different stages of development. As the amount of an 
individual RNA species present at any given time is determined by its net rate 
of synthesis and degradation, an accurate picture of transcription per se can be 
obtained only by directly measuring de novo synthesis of RNA (if you are 
interested in RNA degradation, see Method for measuring mRNA decay rate in 
Saccharomyces cerevisiae). Most techniques employed to measure changes in the 
relative levels of individual transcripts present under different conditions, 
including Northern analysis (see Northern blotting), RT-PCR (see 
Reverse-transcription PCR (RT-PCR)), nuclease protection assays (see Explanatory 
Chapter: Nuclease Protection Assays), and genome-wide assays, such as microarray 
analysis and high throughput RNA sequencing, measure changes in the steady-state 
level of a transcript, which may or may not reflect the actual changes in 
transcription of the gene. Recent studies carried out in fission yeast have 
demonstrated that increases in the steady-state level (accumulation) of many 
individual mRNAs occur without any significant changes in transcription rates 
(McPheeters et al., 2009), highlighting the important role of regulated RNA 
stability in determining gene expression programs (Harigaya et al., 2006).