Indirect Fluorescent Labeling of Microarray Targets Using ImProm-II™ Reverse Transcriptase

Historically, fluorescent cDNA targets used for microarray hybridization have been produced using direct incorporation of nucleotides coupled to bulky fluorescent dyes such as Cy®3 and Cy®5. These modified nucleotides are not efficiently incorporated by reverse transcriptases. Moreover, reverse transcriptases incorporate different dye-modified dNTPs at different rates. Recently, indirect labeling of cDNA target populations, using aminoallyl-derivatized dUTP incorporation followed by amino-coupling of fluorescent dyes, has been gaining ground on the traditional method of direct incorporation.

Promega Notes 81, 14–15.