Dear All,
First I would like to thank you all for this wonderful forum. Very useful information indeed.
I am currently preparing Illumina libraries using Ethan's protocol (http://ethanomics.wordpress.com/chip...useq-adapters/).
The starting cells' number was around 400K. So I amplified the Genomic DNA obtained from ChIP experiment using Adli et al., 2011 protocol (http://www.nature.com/nprot/journal/....2011.402.html)
After the genomic DNA amplification procedure ends, I start with Ethan's protocol from the End repair step. I use 5 cycles for converting Y shaped adaptors. I excise the gel at 200-500 bp. Then I amplify the library using 11 cycles.
At the end I check the QC using agilent High sensitivity Chip (1:20 dilution). Some of the samples show good quality electropherogram while some others show noisy, pointy (spiky) and weird peaks. The range of the peak is correct but it has a weird appearance. I have ran all the samples that shows such peaks in a High sensitivity Bioanalyzer Chip (1:20) and have attached the PDF report of that run.
Have anyone seen such kind of peaks before after ChIP-seq library preparation? Can we sequence such libraries? What causes such appearance of peaks (as in the PDF report)?
I would really appreciate all your help and valuable suggestions...
With warmest regards,
Chadi
First I would like to thank you all for this wonderful forum. Very useful information indeed.
I am currently preparing Illumina libraries using Ethan's protocol (http://ethanomics.wordpress.com/chip...useq-adapters/).
The starting cells' number was around 400K. So I amplified the Genomic DNA obtained from ChIP experiment using Adli et al., 2011 protocol (http://www.nature.com/nprot/journal/....2011.402.html)
After the genomic DNA amplification procedure ends, I start with Ethan's protocol from the End repair step. I use 5 cycles for converting Y shaped adaptors. I excise the gel at 200-500 bp. Then I amplify the library using 11 cycles.
At the end I check the QC using agilent High sensitivity Chip (1:20 dilution). Some of the samples show good quality electropherogram while some others show noisy, pointy (spiky) and weird peaks. The range of the peak is correct but it has a weird appearance. I have ran all the samples that shows such peaks in a High sensitivity Bioanalyzer Chip (1:20) and have attached the PDF report of that run.
Have anyone seen such kind of peaks before after ChIP-seq library preparation? Can we sequence such libraries? What causes such appearance of peaks (as in the PDF report)?
I would really appreciate all your help and valuable suggestions...
With warmest regards,
Chadi
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