The reverse transcription of mRNA into cDNA is the first key step in the study of gene expression. Depending on the downstream application, cDNA synthesis and analysis can be performed as a one-step RT-PCR (where the cDNA is generated and PCR amplified all in one reaction), or a two-step RT-PCR (where cDNA is generated and multiple separate PCR reactions can be performed). To facilitate this, there are numerous systems commercially available for both types of RT-PCR. The SuperScript® III enzyme is the latest version of the reverse transcriptase from Invitrogen, with higher thermostability and longer half-life than SuperScript® II. A number of formats are available for SuperScript® III in kit format, the most recent being the SuperScript® III First-Strand Synthesis SuperMix. In this system, the individual components of a first-strand synthesis kit are combined into two master mixes.
I have used a number of reverse transcriptases, including SuperScript® I, II and III in a variety of formats, most often the SuperScript® II/III First Strand Synthesis System. After RNA/mRNA isolation from cells/tissue, I have used cDNA generated with SuperScript® enzymes for qPCR of multiple genes, 5’/3’ RACE and cloning of full length cDNA. With the First Strand Synthesis System, I have successfully performed all of these applications. The principal reason for using the SuperMix is to replace the individual addition dNTPs, DTT, SuperScript®, RNase inhibitors, Magnesium Chloride and RT Buffer with only having to pipette from two SuperMix tubes which collectively contain all of these components. Using the SuperMix is both time-saving and easier.
I found the SuperMix format particularly useful after generating RNA from experiments with multiple conditions where cDNA was needed for a number of different genes for analysis by qPCR. This enabled a quicker approach to setting up the multiple RT reactions. A particular improvement of Superscript III over previous versions is the ability to perform the RT reaction at higher temperatures than conventional RT enzymes. This can improve the yield of difficult templates with RNA secondary structure (as I found), which would not work with other RT enzymes.
In conclusion, I would recommend using SuperScript® III Supermix for RT-PCR if you want a quick and easy format which results in good cDNA yield.