The QualTrace III module in PeakTrace 6 can generate text based Trace Report files on each trace file basecalled by PeakTrace. These files can be supplied to final customers to help them understand why their trace is not to the quality level they might wish and also what they might do to correct the problems.

The Trace Report files can be generated with increasing levels of report detail, depending on the user needs. The report files are named after the trace file with “_report.txt” added (e.g. if the trace file name is “trace_abc.ab1” then the trace report file will be named “trace_abc_report.txt”) and saved to the same folder as the output trace file.

The Trace Reports can be generated with increasing levels of detail as desired. Trace Reports are only available with the PeakTrace:Box version of the PeakTrace Basecaller and are not available with the Auto PeakTrace RP system. The details of what each output means are described in the QualTrace III User manual.

Auto PeakTrace 6.50 introduced QualTrace III only trace reports, allowing them to function independently of PeakTrace. When qualtrace is selected as the basecaller, all references to PeakTrace (e.g. PeakTrace class, PT Good Bases, and Improvement) are omitted from the trace report. In addition, the PeakTrace settings section will not be included in the full report.

The four Trace Report options and outputs are:

No Report

This option prevents the output of a trace report. This is the default value.

Brief

Outputs a short description of the most important QualTrace III analysis results. An example is shown below.

SUMMARY
Trace File: ABI_3130_1200_B.ab1
PeakTrace Class: Success
Total Bases: 1367
KB Good Bases: 1333
PT Good Bases: 1340
Improvement: 0.5%
Date: 2015/08/11 10:55:00
Trace Score: 50
Contiguous Read Length: 1344

Standard

Outputs a summary and detailed description of the important QualTrace III analysis results. An example is shown below.

SUMMARY
Trace File: ABI_3130_1200_B.ab1
PeakTrace Class: Success
Total Bases: 1367
KB Good Bases: 1333
PT Good Bases: 1340
Improvement: 0.5%
Date: 2015/08/11 10:57:38
DETAILS
QualTrace Class: Excellent trace
Average Peak Spacing: 14.41
Run Time: 9000 sec.
Total Scans: 17960
Detector Overload: 0
Trace Score: 50
Contiguous Read Length: 1344
Dye Signal: 0.94
Bubble Spike(s): 0
Void Peak(s): 0
First Peak at Scan: 1276
Early Signal to Noise: 80
Mid Signal to Noise: 64
Late Signal to Noise: 46
Signal at Noise Level at Base: 1406
InDel(s): 0
Early Mixed Peaks: 0.00
Late Mixed Peaks:0.00
Peak Width at Base 700: 13.29

Full

Outputs the summary and full details of the QualTrace III analysis in text form including the PeakTrace and QualTrace III settings used for the analysis. An example is shown below.

SUMMARY
Trace File: ABI_3130_1200_B.ab1
PeakTrace Class: Success
PeakTrace Code: 0
Total Bases: 1367
KB Good Bases: 1333
PT Good Bases: 1340
Improvement: 0.5%
Date: 2015/08/11 11:00:21
DETAILS
QualTrace Class: Excellent trace
QualTrace Code: 40
Sequencer: 3730-16112-027-16112-027
Run Module: XLRSeq50_POP7_1
Run: Run_3730-16112-027_2007-03-17_16-00_0297
Run Date: 2007/03/17 17:00:52
Original Basecaller: PT 5.51
Capillary Number: 23
Capillary Length: 50
Polymer Lot: 0701015
Tray: 343
Tray Well: B5
Sample ID: 58e5d004d3f311ebaabf000d56d7a0f8
Sample Name: 23997
Sample Comment: <ID:23997>
Sample Owner: Daniel_Tillett
Sequencer User: Daniel_Tillett
Run Temperature Set: 60 deg.C
Run Voltage Set: 6.10 kV
Scan Rate: 0.25 sec.
Measured Run Voltage: 6.0 kV
Measured Run Current: 143.0 mA
Measured Run Power: 25.0 mW
Measured Run Temperature: 60.0 deg.C
Injection Voltage: 15.0 kV
Injection Time: 15 sec.
Average Peak Spacing: 14.41
Run Time: 9000 sec.
Total Scans: 17960
Detector Overload: 0
Trace Score: 50
Contiguous Read Length: 1344
Average Detector Noise: 5.21
Spectral Calibration: 0.08
Dye Signal: 0.94
Bubble Spike(s): 0
Void Peak(s): 0
First Peak at Scan: 1276
Early Signal to Noise: 80
Mid Signal to Noise: 64
Late Signal to Noise: 46
Input Total Bases: 1406
Signal at Noise Level at Base: 1406
Quality Score Threshold: 20
InDel(s): 0
Early Mixed Peaks: 0.00
Late Mixed Peaks: 0.00
Peak Width at Base 700: 13.29

PEAKTRACE SETTINGS
Trace File Output: ab1
Seq File Output: abi
.phd.1 file output: no phd
Good Base Improvement: -20
Good Quality Threshold: 20
N Base Threshold: 5
Mixed Peak Threshold: 0
Signal Start Peak: auto
Basecaller: peaktrace
Fuse Trace at Base: 20
Q Average Trim: 12 20
Force Processing: no
True Profile: no
Skip Short Traces: no
Trim Improved Traces: yes
Stealth Mode: no
Noise Raw Data: no
No Peak Resolution: no
Clean Baseline: no
Extra Normalization: no

QUALTRACE SETTINGS
Delayed First Peak: 133%
Crosstalk Threshold: 15%
Signal to Noise Base: 700
Signal to Noise Threshold: 4
Early Signal Base: 80
Mid Signal Base: 64
Late Signal Base: 46
Poor Resolution Threshold: 300%
Poor Resolution Base: 700
Quality Threshold: 20
Contiguous Readlength: 20 20
Mixed Peak Threshold: 50%
Mixed Trace Threshold: 40%
Early Mixed Base Start: 50
Early Mixed Base End: 250
Late Mixed Base Start: 500
Late Mixed Base End: 700
Bubble Spike Threshold: 200%
Void Peak Threshold: 300%