UV crosslinking of RNA to nylon membrane enhances hybridization signals

Abstract

An improvement in the detection by nucleic acid hybridization of size-fractionated RNA immobilized to nylon-based membranes is described. Electrophoretic transfer of RNA to nylon membranes permits a quantitative determination of different RNA transcripts on the same membrane after sequential hybridization using different 32P-labeled DNA probes. UV crosslinking of the RNA to the nylon membrane increased the intensity of the radioactive signals. Using the method reported here, increased signals of between 10 and 40 fold were observed, depending on the species of transcript tested. Moderately abundant as well as rare transcripts can easily be detected in as little as 5 micrograms total cellular RNA.