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Hi everybody,
this might be a stupid question but I did not get it. Why is the size selection step in the ChIP library prep process prior PCR enrichment?
I suppose using the whole amount of adapter ligated library will result in a more complex library with less duplicates. Size selection (I use the Pippin) can be done after PCR enrichment.
But there must be a point as other protocols also perform the size selection step pre PCR.
I thought about residual adapter from the ligation step but RNA or DNA lib prep enrichment is not perturbed by this.
Thank you very much for your experiences and ideas,
Elisabeth
Just as additional information: I use Illumina's TruSeq ChIP seq lib prep Kit. After adapter ligation there is the standard AMPureXP clean up step followed by size selection (I perform using the Pippin prep). After this I do an additional AMPureXP step to reduce the Pippin elution volume from 40 to 20 µl. After this I do the PCR step.
this might be a stupid question but I did not get it. Why is the size selection step in the ChIP library prep process prior PCR enrichment?
I suppose using the whole amount of adapter ligated library will result in a more complex library with less duplicates. Size selection (I use the Pippin) can be done after PCR enrichment.
But there must be a point as other protocols also perform the size selection step pre PCR.
I thought about residual adapter from the ligation step but RNA or DNA lib prep enrichment is not perturbed by this.
Thank you very much for your experiences and ideas,
Elisabeth
Just as additional information: I use Illumina's TruSeq ChIP seq lib prep Kit. After adapter ligation there is the standard AMPureXP clean up step followed by size selection (I perform using the Pippin prep). After this I do an additional AMPureXP step to reduce the Pippin elution volume from 40 to 20 µl. After this I do the PCR step.