Primer extension is often used to map the 5′ end of RNA (1 ). A single-stranded, end-labeled DNA primer is hybridized to RNA first. Using an RNA-dependent DNA polymerase (reverse transcriptase [RT]) and nonradioactive deoxynucleotides, the primer is extended to yield cDNA. The cDNA is then analyzed on a sequencing gel to determine nucleotide length. The length of the cDNA reflects the distance between the primer and the 5′ end of RNA, and hence, maps the transcription start sites.