Advantages of StaRT-PCR over real-time include a) the data are numerical and standardized, b) quantification takes place at endpoint of PCR so expensive equipment and sequence specific fluorescent probes are not necessary, c) the reactions can be done in multiplex (at least 96 in one reaction) which saves on cDNA. The data reported here were done in multiplex, but we also measured the System 1 genes (over 90 of them) in multiplex. The reproducibility between data obtained in uniplex and multiplex was excellent. Because the data reported here are standardized, all laboratories should obtain the same results we did using standardized CT mixtures Systems 1-3 from Gene Express National Enterprises (G.E.N.E.) Inc.

Listed are the 278 genes that may be measured using G.E.N.E., Inc. Systems 1-3 reagents for StaRT-PCR. Thus far, we have measured 242/278, and the values are listed in descending order in units of mRNA/10E6 b-actin mRNA. The genes with a value of zero are those for which there was no native template product observed when only 6 molecules of the competitive template were present in the reaction. The genes represented in Systems 1-3 that we have not yet attempted to measure are listed at the end.

For most genes, triplicate values were obtained and the value provided is the mean. There are a few for which we have obtained only one value.

For this study, the average CV for genes with at least triplicate measurements was less than 0.25.

Validation of StaRT-PCR includes a multi-laboratory blinded study described in a manuscript to be published in Molecular Diagnosis in December. I would be happy to provide a pre-print to those interested.