RT-qPCR Detection of Senescence-Associated Circular RNAs
Primary cells that reach the end of their replicative potential, encounter sublethal stress, or experience the activation of certain oncogenes cease proliferation and enter a state of long-term growth arrest named senescence. The senescent process has been implicated in a variety of age-related diseases and also in the pathogenesis of cancer. Senescence is characterized by distinct changes in the types and levels of coding RNAs (mRNAs) as well as in the vast collective of regulatory noncoding (nc)RNAs, which includes microRNAs, long noncoding RNAs (lncRNAs), and circular (circRNAs). Numerous technologies permit the detection of senescence-associated linear transcripts (mRNAs, lncRNAs, microRNAs), but the identification and quantification of circRNAs in senescence require distinct molecular approaches. In this chapter, we describe a method for the detection and measurement of circRNAs in senescent cells using specialized reverse transcription (RT) followed by real-time quantitative (q)PCR analysis.
Key wordsRNA-binding proteins Sponge circRNAs Divergent primer design Transcriptome
This work was supported in its entirety by the National Institute on Aging Intramural Research Program of the National Institutes of Health.
- 2.Memczak S, Jens M, Elefsinioti A, Torti F, Krueger J, Rybak A, Maier L, Mackowiak SD, Gregersen LH, Munschauer M, Loewer A, Ziebold U, Landthaler M, Kocks C, le Noble F, Rajewsky N (2013) Circular RNAs are a large class of animal RNAs with regulatory potency. Nature 495:333–338CrossRefPubMedGoogle Scholar