To be skewed or not to be? This is no longer the question

摘要

In their letter to the editor, Dr Prchal's group raises several issues. We agree with some of these issues and respectfully disagree with others. First, the T_m for the 3 IDS primers used by both our groups varies between 51.6degC and 52.9degC (not 60degC). Using annealing temperatures between 45degC and 55degC, we observed identical Ct values and excellent discrimination between IDS alleles, whereas an annealing temperature of 60degC resulted in less efficient amplification. Furthermore, we obtained identical results using (he carboxy-fluorescein-labeled /DS-specific probe (used by Swierczek et al1) and SYBR green. Thus, as reported,2 the quantitative allele-specific polymerase chain reaction transcription assay (TCA) that we used was not identical, but very similar, to that used by Swierczek et al.