A modified method for the detection of differentially expressed mRNAs without using radioactivity.

摘要

We present here a modification of the original differential display approach using a single oligo(dT) primer for the reverse transcription reaction (instead of the various oligo(dT)NM primers that subdivide the pool of mRNAs) and a combination of 25-mer or 26-mer arbitrary primers together with 30-mer anchored primers for the PCR reaction. The PCR products are, then, efficiently separated in a non-denaturing polyacrylamide gel and the bands are visualized after staining with silver nitrate. The model for the development of our differential display approach was seven clones of an insect species: the aphid Myzus pesicae (Sulzer) (Homoptera: Aphididae). We believe that our modified differential display technique, with the efficient resolution of the DNA bands in a non-denaturing gel and staining with silver can be applied as an alternative non-radioactive detection of differentially expressed messages in various cell populations. In addition, the method could be used as a supplementary tool to other techniques for examining inter- and intraspecific genetic variation in aphids.