Primer dimer problem

[cybeline]

If you sequence it like this, you will lose a lot of data due to the machine just sequencing primer dimers.
You could size select using AMPure beads. That will give you better yield than gel extraction.
When you do your ligations next time, use less adapter and be sure to add the ligase last to your reactions, not in a master mix.

Thank you very much for your answer.

I will look into Ampure beads for the cleanup.
But I believe the primer diner occurs during the pcr enrichment step since after the ligation I gel purify my samples.

[pbluescript]

If you sequence it like this, you will lose a lot of data due to the machine just sequencing primer dimers.
You could size select using AMPure beads. That will give you better yield than gel extraction.
When you do your ligations next time, use less adapter and be sure to add the ligase last to your reactions, not in a master mix.

[cybeline]

Hello,

I have just finished prepping multiplex samples for paired end Illumina sequencing using NEB products but after a bioanalyzer I have a lot of primer dimer at 75bp and around 120bp which seems even stronger than my bands at 300bp.
The sequencing facility does not want to sequence them due to the primer dimer problem.

Anybody else having the same problem?

I was suggested to re-gel purify all the samples but I am afraid that I will end up with not enough DNA to sequence.

Any other suggestion or the effect it could have if we sequence it like this?

Attached is a picture of the caliper data.

Thank you,