Tech Spotlight: Environmentally-friendly, High-density Fluorescent PCR Labeling Techniques for Nucleic Acids

Many molecular biology techniques rely on the ability to label DNA and RNA with detectable markers. CU researchers led by Robert Kuchta have developed fluorescent cytosine triphosphate (dCTP) analogues for use in PCR and subsequent DNA analysis. This technology improves the polymerase chain reaction (PCR) by generating simple, high quality, immediate visualization of PCR products, and eliminates the conventionally required staining step involving toxic ethidium bromide. Analogue compounds are incorporated into products through a modified PCR technique with a higher denaturing temperature and readily available DNA polymerase.

This research group has also developed a method for the synthesis of fluorescently labeled RNA based on the incorporation of the fluorescent cytidine analogue tCTP by T7 RNA polymerase in transcription. This methodology can be used to directly detect RNA without the use of radioactivity or staining, and can also be used to generate fluorescent aptamers for detecting specific analytes.

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