Sequencing data with three spikes
Raw data view showing same three spikes. This view is only available using ABI Prism Sequencing Analysis.
Occasionally a spike occurs in the data and the base caller software, either ABI Prism Sequencing Analysis or PHRED, calls the spike an "N", or an incorrect base upon interpreting the raw data. Spikes are the result of dust or sieving matrix (polymer) crystals in the capillary reflecting the laser back onto the detector. We keep our machines clean and inspect the quality of the polymer as best as possible to prevent this. The laser light contains the emission spectra of all four dyes and for this reason the spikes consist of all four colors. Fortunately these spikes are only on the order of two scans wide and are easily discernable from the surrounding fifteen scan wide peaks. In the sequencing data above the sequence surrounding the two spike areas are . . . TAT(C)ACC . . . and . . . AAGA(N)TACC where the bases in parenthesis are the incorrect calls. Through viewing of the raw data we can usually edit the sequence data. It is now easy to see that the actual sequence is . . . TATgACC . . . and . . . AAGAaTACC. This is a typical case as far as spikes are concerned however, when your data's signal is weak, the spikes show up more and for that reason are a greater nuisance. We no longer automatically edit these spikes since our clients use the data for numerous reasons. We will edit them upon request and return them via email. Any that we can not confidently call, we will reinject and return via DNALIMS.