The skip short traces allows PeakTrace 6™ to skip over traces that have less than a certain number of bases and which may not be improved by PeakTrace. This setting is useful because the KB Basecaller typically basecalls short read DNA sequence well and hence the improvement from using PeakTrace can be minimal. By using this setting users can avoid having to manually preselect the traces for PeakTrace basecalling, a time consuming and often inaccurate step.
This function is aimed at users who are only interested in improving longer runs, or would like to avoid having to manually preselect the traces that will see the most improvement from PeakTrace. The reads below the set read length are skipped over and the cost of using PeakTrace avoided on these traces.
When using this function, trim settings, noise raw data and stealth mode will all still be applied to the short trace at no charge (Figures 1 & 2). If it is desired that trimming is not performed on the short traces then the trim improved traces only function should be selected. Trim improved traces only will apply the trimming to the PeakTrace processed traces only and not to the short traces skipped over.
Figure 1. KB basecalled short trace before processing
Figure 2. Trace skipped by Auto PeakTrace 6. Note that this trace has had trimming applied, however the rest of the trace is identical to the original file
Potential Pitfalls with using Skip Short Traces
If the skip short/pcr base option is set above 500 bases then traces that could have been improved will be charged. A setting lower than 500 will allow more traces to be processed and the functionality of the skip short traces option will remain on, however a setting of 0 in the skip short/pcr base option will effectively turn skip short traces off, as processing will be attempted for all traces with more than 0 bases.
If this setting is used with those settings that improve the KB traces (i.e. clean baseline, extra normalization, and extra baseline)then PeakTrace will improve the KB trace and this will become a charged trace. To avoid this, set the basecaller to peaktrace and/or unselect any of KB improving settings.