Weird Illumina library

[ashchin]

Hi ,

Has anyone observed a picture like this (please find the link below) after running the illumina library on Bioanalyzer.
Any ideas what would have caused this?? I was expected an ideal 350 bp peak.

http://imageshack.us/photo/my-images/189/illuminalibrary.png/

Thank you

[Heisman]

Your tallest peaks are all roughly 55-60bp apart, so maybe adapter dimer laddering of some sort?

[Jon_Keats]

What kind of library? I've seen some odd curves on RNAseq libraries prepped from degraded RNA.

[ashchin]

Thank you, this is a DNA sample (FFPE).
It looked fine after fragmentation but after library prep it looked weird.

[ashchin]

@Heisman - thats what I thought it was, also they have a 30 bp difference between peaks.

[ETHANol]

I've had the bioanalyzer give strange banding patterns before and rerun the same sample and had it look fine. That being said, did you use the TruSeq In-line controls (I hope not!)? Perhaps it is the in-line control DNA.

[ashchin]

I have not used the in line controls. It actually looked fine the first time I ran on the bioanalyzer. My qpcr failed twice so I re ran my sample and this is hoe it looked, which is really strange.

[madseq]

We once got a library with 30 bp difference between peaks, which was due to adapter concatemers. We ended up sequencing the adapters. We now know that this concatemerization phenomenon can occur if the starting material is very low, but it can be prevented by lowering the amount of adapters at the ligation step.