Does this band make sense? Limits to expression based cancer studies

摘要: Cancer researchers commonly employ reverse transcriptase polymerase chain reaction (RT-PCR) for gene expression analysis of cancer cells. While this technique is facile and reproducible, it not without limitations. The human genome contains abundant nearly identical sequences (e.g. pseudogenes) to mRNA transcript sequences, which amplify when performing RT-PCR on samples with even trace amounts genomic DNA. Such include housekeeping transcripts such as beta-actin GAPDH. This also true numerous products whose altered in disease states pp32). Moreover, we describe that amplification undesirable simply avoided by designing primers spanning multiple exons. We found template-specific reactions lack the specificity necessary definitively determine sense or anti-sense orientation an transcript. Given above mentioned caveats limitations studies, encourage investigators test existence intronless are similar specific being studied. Further, RNA should be completely DNA-free prior based assays. Finally, ensure reliability array results, recommend utilizing widely accepted loading controls, GAPDH and/or beta-actin.