Measuring the length of poly(A) tails.

Adenylation status has an important role in the regulation of mRNA metabolism: mRNAs are deadenylated before degradation, microRNAs (miRNAs) can cause deadenylation, and the poly(A) length of certain mRNAs is regulated during development. This protocol describes methods that can be used to measure the poly(A) tail length of specific mRNAs. These include, in the order of increasing sensitivity, (1) northern blotting of intact and experimentally deadenylated mRNAs and (2) northern blotting of intact and experimentally deadenylated mRNA fragments that have been cleaved near the 3' end with RNase H. Highly sensitive polymerase chain reaction (PCR)-based approaches are also discussed.