I also noticed this phenomenon with TruSeq RNA libraries. (See here.)
(Note this should not be confused with the post-enrichment-amplification double peak phenomenon. This phenomenon, when we see it, occurs prior to enrichment-amplification.
I have a couple of hypotheses as to what might be going on. What is your take?
--
Phillip
(Note this should not be confused with the post-enrichment-amplification double peak phenomenon. This phenomenon, when we see it, occurs prior to enrichment-amplification.
- Below two genomic DNA samples (~1ug each) were Covaris S2-fragmented, then ampure concentrated/shaped to remove the smallest fragments and any contaminating junk from the DNA prep.
- Each was split into 2 replicates, with each of the replicates being carried through the rest of the protocol. (Different adapter index used on each replicate.)
- The legend tells the story. Peak initially at ~200 bp after blunting/a-tailing.
- After ligation the main peak appears to be converted into a double peak. Or, in the case of the second set of libraries, a main peak with a larger shoulder.
- Following 6 cycles of enrichment PCR only the right-most peak appears to be retained.
I have a couple of hypotheses as to what might be going on. What is your take?
--
Phillip
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