A Quick, Affordable Method for the Quantification of Gene Isoforms

Most genes are transcribed as several variants known as isoforms. Isoforms are transcripts from the same locus with either a different transcription start site or post‐transcriptional modifications, for example alternative splicing. As a result, isoforms might be characterized by different functions and/or stability. The differences between gene isoforms span from a few bases to several kilobases. In some cases, no region of the transcript might be specific to only one isoform, making quantification problematic. We devised a quick, reliable, one‐step method for the accurate quantification of gene isoforms, even in the latter case. We compared the isoforms' sequences using bioinformatic tools freely available online. We analyzed RNA samples with a one‐step real‐time RT‐PCR protocol, using primer sets specific to one or more isoforms and RNA standard curves as a reference. Differential analysis of the data provided reliable information on the copy number of each isoform in the original sample. This method might provide a first step in understanding the importance of differential expression of gene isoforms This work was supported by a D'Youville College research grant.