Reverse transcription-polymerase chain reaction: an overview of the technique and its applications

The polymerase chain reaction (PCR) has galvanized molecular biologists by virtue of its ability to provide them with large quantities of any desired fragment (up to 11kb) of DNA. This power combined with its flexibility has also inspired many useful applications, including new methods of DNA sequencing, cloning and mutagenesis. One logical variation of PCR has been its application to the detection and analysis of messenger RNA by the addition of a reverse transcription step prior to performing PCR. Due to the exquisite sensitivity of PCR, reverse transcription-PCR (RT-PCR) has been used to characterize mRNAs previously undetectable by established methods of RNA analysis such as Northern hybridization and RNase protection assays. Furthermore, its capacity as a method of quantitative analysis is currently being developed. RT-PCR has also been used to diagnose the presence of certain diseases. Recently, RT-PCR has been employed to identify and isolate genes that are differentially expressed in different cells or environmental conditions.