Table 3.
PCR Reaction Mix for fully automated detection of Mit1C and IAC using PerfeCTa or PCRBio master mix.
| Component * | Final Concentration |
|---|---|
| Nuclease-free water | Variable |
| PerfeCTa Multiplex 5× or PCRBio 4×/dUTP/UNG | 1× |
| Mit1C primer (Forward) | 400 nM |
| Mit1C primer (Reverse) | 400 nM |
| IAC DNA (500 copies/µL) | 1.5 µL |
| IAC primer (Forward) | 400 nM |
| IAC primer (Reverse) | 400 nM |
| Template | Variable |
| Total Volume | 15 µL |
* The probes in the Rheonix Assay are spotted on the filter. (a) PerfeCTa multiplex qPCR Toughmix Low ROX, Quantabio LLC., Beverly, MA, USA, Cat# 95149-250 and (b) PCRBio master mix (PCR Biosystems, London, UK, #PB25.53-P4) with UNG (ArcticZymes, Tromso, Norway, Cat#75011) and dUTP (Promega, Madison, WI, USA, Cat# U1191).