Sharp peaks in the trace signal
Identification of sharp sequencing trace spikes
- The trace file contains sharp spikes (from 1 - 2 to 8 - 9 data points wide) in the raw data channel that can have intensities as high as 32,000 (detector limit).
- The sharp peaks is normally higher than the surrounding sequence peaks.
- The sharp peaks are generally more common towards the end of the trace than the beginning but they can occur anywhere in the trace.
Figure 1. Example of a small sharp signal spike. (A) KB processed data channel data. (B) Raw channel data at the same location.
Figure 2. Large signal spike (raw channel). Note that the signal reaches an intensity over 32,000.
Cause of sharp trace signal spikes
- The cause of sharp signal spikes is a mystery. The current best theory is these peaks are caused by small gas bubbles forming in the capillaries during electrophoreses which refract the laser light into the CCD camera when they pass by the detector zone. The frequencies of sharp spikes seems to be operator and machine dependent.
Solving sharp trace signal spikes
- The best solution is to purge buffers and load fresh matrix. Avoiding excessive shaking of the buffers may also help.
Return to the main DNA sequencing troubleshooting page.