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The FairPlay system uses a two-step chemical coupling method to fluorescently label cDNA. First, an amino allyl-dNTP is incorporated into the first-strand cDNA. For this step, the kit includes AffinityScript HC Reverse Transciptase, our newly engineered thermostable mutant of MMLV reverse transcriptase that exhibits enhanced RNA template binding capability. This improved reverse transcriptase allows you to produce full-length cDNA and also efficiently reverse transcribe on RNAs with secondary structure. The amino allyl-dNTP does not exhibit steric hindrance in the reverse transcription reaction. As a result, cDNA is amino-modified with amino allyl-dNTP uniformly incorporated at a higher frequency than a directly labeled Cy™ dye dNTP. Second, an amine-reactive Cy dye is chemically coupled to the amino-modified cDNA. The FairPlay system efficiently creates evenly labeled cDNA of high fluorescence intensity without any dye bias.

Greater Sensitivity Means Less RNA
The FairPlay labeling method is very robust, allowing you to use as little as 5 to 10 µg of tota RNAl or 200 to 500 ng of poly(A) RNA. The high rate of incorporation of the amino allyl-dNTP to create amino-modified cDNA along with efficient coupling of the amino-modified cDNA to the amine-reactive fluorescent dyes, makes the FairPlay labeling method highly sensitive and allows detection of low abundance genes. With the same amount of starting material as other kits, the FairPlay II kit gives you a better signal. You can use as little as 2 µg of total RNA to detect signal; however with more RNA the signal is enhanced.