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I have sequenced numerous multiplexed pools of BS amplicon-seq libraries derived from human samples on a MiSeq over the past few weeks. I have been utilising trim-galore and Bismark for alignment and am finding the mapping efficiency to be really low for two pools (55% & 30% respectively) both of which had a cytosine per base sequence of around 10-20% throughout the entire read in their fastqc files.
The high C content visible in the fastqc files makes me think the poor mapping efficiency is due to poor bisulfite conversion, as this would be expected to be close to zero is bisulfite conversion had actually took place.
I am new to the field so any help would be greatly appreciated.