An improved primer extension method for detection of mRNA start-points using non-radioactive digoxigenin-labeling primers.

An improved primer extension method using non-radioactive digoxigenin (DIG)-labeling primers is described which uses a commercially-available DIG-labeling and detection system to perform alternative hybridization-based "labeling" procedure of DNA markers and DIG chemiluminescent detection assay. The time-consuming annealing step and relatively low specificity of conventional protocol are also improved considerably by an application of one-step primer/mRNA annealing procedure and subsequent high-temperature reverse transcription reaction. This new protocol is convenient, simple, cost-effective and safe, and can allow the detection of even low abundance mRNA start-points in enriched poly(A)(+) RNA samples.