Sample And Data Troubleshooting - Thermo Scientific Applied Biosystems SeqStudio Genetic Analyzer User Manual

Sample and data troubleshooting

Observation
Offscale signal is detected is
displayed
Note: Samples are stable on
the instrument for 16–24 hours.
Determine if samples will be
stable if a re-injection is
recommended, then plan the
re-injection accordingly.
There is no signal or a low
signal
Note: Samples are stable on
the instrument for 16–24 hours.
Determine if samples will be
stable if a re-injection is
recommended, the plan the re-
injection accordingly.
SeqStudio Genetic Analyzer Instrument and Software User Guide
™
See also "Data quality alerts" on page 119, "Sizecalling and basecalling quality
alerts" on page 119, and Troubleshooting Sanger sequencing data User Bulletin
(Pub. No. MAN0014435).
Possible cause
The sample concentration is
too high.
The injection conditions are too
strong for the sample.
The sample volume was
insufficient.
The sample concentration was
too low.
There were bubbles in the
sample wells.
The sequence reaction failed.
The Hi‑Di Formamide used to
™
prepare the samples was
degraded.
The sample was prepared with
the BigDye XTerminator
Purification Kit but a BDX run
module was not selected.
The sample was degraded.

Sample and data troubleshooting

Recommended action
Dilute the sample, prepare a new plate, and
start a new run.
Quantitate the sample prior to adding reagents
for capillary electrophoresis.
Re-inject the samples with adjusted injection
conditions.
Injection time, voltage, or a combination of
these conditions can be adjusted.
Use the recommended sample volume.
See "Sample preparation guidelines" on
page 37.
Re-inject the samples with adjusted injection
conditions.
Injection time, voltage, or a combination of
these conditions can be adjusted.
Centrifuge the sample plate or tubes to remove
the bubbles before loading onto the
instrument.
If the samples have been run, centrifuge the
plate or tubes, then set up a re-injection.
Review the sequence analysis protocol, the
template quality, and the template quantity. Set
up a new plate and repeat the reaction.
Troubleshooting Sanger sequencing data
See
User Bulletin (Pub. No. MAN0014435).
Prepare the samples with fresh Hi‑Di
Formamide and repeat the experiment.
See "Sample preparation guidelines" on
page 37.
Re-inject the sample with the correct module.
™
Prepare the sample according to the protocol
provided with the kits for sample preparation.
See "Sample preparation guidelines" on
page 37 for sample preparation guidelines.
IMPORTANT!
water.
Appendix A Troubleshooting
™
Do not resuspend samples in
A
175