What is the adapted annealing protocol for PyroMark Q24 Advanced procedure?

The sequencing reaction volume was reduced to 20µl. The amount of sequencing primer diluted in annealing buffer is unchanged so that the sequencing primer concentration is increased. After releasing the beads into the sequencing reaction, the plate is heated at 80°C for 5 min and directly loaded into the instrument (versus standard PyroMark Q24 procedure 80°C, 2 min followed by 5-15 min incubation at room temperature). The advanced run is started directly and always begins with 10 min of shaking before dispensing the enzyme mix.