7500 fast system sds

In addition, the samples (n=30, except the 7 excluded samples) were finally analyzed using the L1, MGP5+/6+ primer system 11 (link). This was performed to further investigate possible reasons for conflicting results in the reinvestigation process and to examine the negative samples using a general method. Analyses were carried out in sample duplicates of 25 μL together with negative and positive controls for both MGP and the human control gene HBB. Sample reactions contained 1x Quantitect SYBR green master mix (Qiagen, Germany), either 0.3 μM forward and reverse primers for HBB or 0.3 μM forward and reverse MGP primer mix and DNA of ∼50 ng, and were manually applied on 96-well plates. Subsequent analysis was performed on the 7500 fast real-time PCR system (Applied Biosystems, the Netherlands). The standard program was used with a denaturation step at +95°C for 10 min, followed by 5 cycles at +95°C for 0.5 min, +42°C for 0.5 min, and +72°C for 0.75 min; 45 cycles at +95°C for 0.5 min, +64°C for 0.5 min, and +72°C for 0.75 min; and a final step at +72°C for 10 min. The software 7500 fast system SDS (Applied Biosystems) was used to analyze results, and the curves were manually assessed using a threshold at 35 cycles for a positive result. Every run was analyzed with a positive and a negative control.