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Updated: May 17, 2026

Adapting 3' Rapid Amplification of CDNA Ends to Map Transcripts in Cancer
Published on: March 28, 2018
Simple and inexpensive three-step rapid amplification of cDNA 5' ends using 5' phosphorylated primers
1Rega Institute for Medical Research, Department of Virology and Chemotherapy, KULeuven-University of Leuven, B-3000 Leuven, Belgium. kaihenrik.dallmeier@rega.kuleuven.be
Abstract:
Rapid amplification of cDNA 5' ends (5'-RACE) is routinely used for the sequence analysis of the upstream noncoding regions of cellular mRNAs; however, it represents a tedious and cost-intensive procedure. By employing 5' phosphorylated gene-specific primers for first-strand cDNA synthesis, we cut short the previously established reverse ligation and amplification protocol of Mandl and coworkers (BioTechniques, 1991, vol. 10, pp. 484-486) to a streamlined three-step procedure that no longer depends on enzymatic mRNA decapping or linker ligation. The novel three-step protocol has been validated by mapping the transcriptional start sites of heterologously expressed yellow fever virus genomic RNAs from cultured mammalian cells.
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