Application Guide - QIAxcel Advanced

QIAxcel Advanced Application Guide 10/2016 7 for capillary electrophoresis with a DNA Screening Cartridge. The AM900 method and QX Alignment Marker 15 bp were used. AM900 is a customized method with the following parameters: alignment marker injection at 4 kV for 20 sec, sample injection at 2 kV for 40 sec, and separation at 3.5 kV for 900 sec. For degradation analyses, the samples were sonicated with 7, 14, 70 and 105 pulses. Ultra-sound degradation was performed in 500 μl (100 ng/μl) of DNA, sonicated on a Branson Sonifier® 250. Results Samples from the dilution series were electrophoresed on a 1% agarose gel (Figure 1A), as well as on the QIAxcel system using a DNA Screening Cartridge (Figure 1B). The results demonstrate that gDNA concentrations between 25 and 100 ng/μl are suitable when using this system. Various running conditions, such as injection time, separation time and voltage, were tested (data not shown) and optimized. AM900 was identified as the optimal procedure for gDNA quality control with sample injections of 40 sec at 2 KV and a separation time of 900 sec at 3.5 KV. The results of assessing sonicated gDNA samples with the QIAxcel system revealed that it is very simple to determine the degree of degradation by categorizing the type of peak yielded from each gDNA sample (Figure 2). A major peak with no signals for degradation products before it and a long tailing off after it indicates gDNA of very good quality that is suitable for downstream applications. Electropherograms for partially and highly degraded gDNA have no tailing off after the major peak. The level of degradation can be differentiated based on the signals for the degradation products in the broad peak before the major peak. A B Figure 1. Electrophoresis of samples from a dilution series. A Lanes 1–5: gDNA samples at respective concentrations of 10, 25, 50, 75, and 100 ng/μl run on a 1% agarose gel for 150 min at 80 V. Lane M: lambda HindIII size marker. B The same samples run using a QIAxcel DNA Screening Cartridge and the AM900 method. M 1 2 3 4 5 M 1 2 3 4 5 M 12345 M 12345

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