Hello,
I've been following the TruSeq DNA sample preparation method to make library preps from 3ug of starting DNA. I am aiming for an insert size of around 600bp. The DNA is sheared using a Covaris instrument and a band is cut out of a 1.2% long agarose gel. After 15 cycles of PCR amplification with longer extension time, the library is cleaned up (with ampure XP beads) and run on an Agilent high sensitivity chip (see picture). We have been getting shoulder peaks of between 300 to 500bp beside the larger main library peak which can affect the final analyses of the results after sequencing.
Has anyone else come across these shoulder peaks and/or been successful in removing them/creating libraries without them?
Thanks for your help,
Anna
I've been following the TruSeq DNA sample preparation method to make library preps from 3ug of starting DNA. I am aiming for an insert size of around 600bp. The DNA is sheared using a Covaris instrument and a band is cut out of a 1.2% long agarose gel. After 15 cycles of PCR amplification with longer extension time, the library is cleaned up (with ampure XP beads) and run on an Agilent high sensitivity chip (see picture). We have been getting shoulder peaks of between 300 to 500bp beside the larger main library peak which can affect the final analyses of the results after sequencing.
Has anyone else come across these shoulder peaks and/or been successful in removing them/creating libraries without them?
Thanks for your help,
Anna
Comment