Bio-Rad SsoAdvanced™ Universal SYBR® Green Supermix - Manual (Page 26)

20 | SsoAdvanced ™ Universal SYBR ® Green Supermix Instruction Manual 20 |
1. Using Table 6, determine the percent of gDNA contamination present. For example, if the
∆Cq (no-RT control Cq – cDNA Cq) for a given sample is seven or greater, then <1% of the
DNA present in the sample is gDNA, which would be considered insignificant.
Table 6. Determining percent of gDNA contamination.
∆Cq
Percent Contribution, %
1 50.00
2 25.00
3 12.50
4 6.25
5 3.125
6 1.5625
7 0.78125
2. Evaluate the assay design and note the location of the primers. To avoid gDNA amplification,
at least one primer must span an exon:exon junction site. Alternatively, the primers can be
designed in two different exons that are separated by an intronic region >1 kb.
If you are using an internal positive control (IPC) and the standard deviation of the Cq values
across all samples is >0.167, then consider the following:
When the IPC for a given sample(s) is higher than the group, this is most likely due to the
presence of a PCR inhibitor. Review the sections on sample preparation for more information.