Fig. 3

SYBR Green is suitable to analyse alternative splicing by RT-qPCR. a Melting curve analysis of the two variant specific fragments amplified with control primers 2 from mRNA transcripts expressed in the choroid plexus of mouse brain in the presence of onefold concentrated (1 ×) SYBR Green or SYBRGreenER. b Amplification curves using a 1:1 mixture of two plasmids containing the cDNA of variants 1 and 2 (1.25 pM each) in the presence of different SYBR Green concentrations. Dye concentrations are indicated as multiples of the concentration recommended by the supplier, since it was impossible to obtain detailed information about the molarity of the dye in the reagent. Dashed lines show the mean of three reactions using control primer pair 1, solid lines using control primer pair 2. c The cycle of quantification (Cq) and the PCR efficiency (E) of the amplification curves shown in C were obtained from the comparative quantitation tool of the Rotorgene software and efficiency corrected Cq values (CqE) were calculated as described [18]. All curves, the Cq and E values are means of triplicates. Experiments were performed thrice with similar result