REVERSE TRANSCRIPTASES
how do they compare
There are many reverse transcriptases available to monitor gene expression. Your selection of reverse transcriptases is one factor that can influence successful real-time and endpoint RT-PCR, labeled-probe generation for microarrays and cDNA cloning. The ideal reverse transcriptase will robustly transcribe the entire length of all RNAs, including the least abundant. The authors of the article Lining Up the Scripts: A Comparison of Reverse Transcriptases performed a direct functional comparison of several popular reverse transcriptases to evaluate:
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A,
B,
Panel A. Human total RNA was reverse transcribed using oligo(dT) according to the manufacturer’s instructions with the exception that the GoScript™ Reverse Transcription reaction was carried out at 50˚C, as recommended for SuperScript® III. cDNA was analyzed by amplification of b-2 macroglobulin (B2M, control) or GC-rich Nocturnin using GoTaq® Green Master Mix (Cat.0x M7122). Panel B. Human total RNA was again reverse transcribed but at each enzyme’s recommended reaction temperature. cDNA was analyzed by real-time PCR for both transcripts using GoTaq® qPCR Master Mix (Cat.0x A6001). Data are the average of three independent experiments.
• erformance when detecting trace P transcripts by real-time PCR using long RNA templates • ensitivity to ethanol, a common S contaminant in RNA preparations • bility to transcribe a challenging A GC-rich mRNA In terms of sensitivity of transcript detection, transcript length and sensitivity to ethanol contamination, the new GoScript™ Reverse Transcriptase performed as well as or better than Invitrogen SuperScript® II and SuperScript® III and Qiagen Omniscript® and Sensiscript® reverse transcriptases. In a comparison of GoScript™
and SuperScript® III using a GC-rich transcript and an elevated reaction temperature (50°C), GoScript™ Reverse Transcriptase outperformed SuperScript® III in the amount of cDNA produced as determined by quantitative PCR. GoScript™ Reverse Transcriptase is more efficient using templates with complex secondary structure. Read more about how GoScript compares in: Lining Up The Scripts: A Comparison of Reverse Transcriptases.
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