LongTrace and poor quality traces

In general LongTrace does not work well with low quality traces. This is because LongTrace improves the signal present and for most low quality traces the signal is either not present (failed reactions, short reads) or can not be made sense of (mixed templates, noise traces).

The major exception to this limitation are traces that have very low resolution (blurry) and traces with high levels of signal crosstalk (large crosstalk look like mixed signal). These types of traces are often improved the most by longtrace. We have seen some traces showing these two problems that have had read length increase of over 150%.

Daniel Tillett
Nucleics Support



Edited 1 time(s). Last edit at 11/13/2006 06:04PM by nucleics.

So what should I do we traces that are not improved by LongTrace?

You really need to look at your sequencing protocol. Our experience is the DNA template seems to be the critical component.

The simplest thing to do is run your traces through our free QualTrace server (http://www.nucleics.com/qualtrace-dna-sequencing-demo/) and see what the it says is the problem with your traces.

If the problem is weak ends then there are two things to try that will help. 1. Try a template concentration range, both higher and lower (10 fold) than what you are currently using, and 2. Try increasing the annealing and extension thermocycling times (15 s and 5 minutes).

Cheers

Daniel Tillett
Nucleics Support