In Vitro Continuation of RNA Synthesis Initiated in Vivo

Isolated nuclei will continue synthesis of RNA initiated in vivo, but reinitiation of synthesis is rare in washed nuclei (1 ). This situation can be exploited to measure instantaneous rates of in vivo transcription because the cell-free conditions are well-defined and nascent transcripts are generally not subject to the rapid cleavage often found in living cells (2 –5 ). Isolated nuclei can also be used to map a primary transcript on genomic DNA. The method has been used to show that transcription terminates more than 1000 nucleotides downstream from the poly A site in (β-major globin mRNA (6 ).