What types of primers are typically used for first strand synthesis in a reverse transcription reaction?
Three options exist for priming a reverse transcription reaction. (1) The classic approach is to utilize oligo(dT) primers. These primers are typically ~20 bases in length, and anneal to the polyA tails of mRNA. By targeting the mRNA fraction, the complexity of the resultant cDNA population is dramatically reduced, since rRNA and tRNA species will not serve as templates in the reaction. The drawback of using oligo(dT) primers is that the resultant cDNA population will have a 3 bias, thus compromising the effectiveness of PCR primers targeting the 5 ends of transcripts. In addition, due to the 3 bias, fragmented samples lacking a polyA tail will not be reverse transcribed. (2) Utilizing random primers is another popular strategy for priming reverse transcription. These are a random mixture of the four DNA bases of a specified oligo length. Random hexamer mixes are commonly used, consisting of 4096 sequences (46). Each of these primers will anneal anywhere the complementary sequence exis
