I am considering using Life's AmpliSeq protocol and modifying it to use Illumina indices for sequencing on a NextSeq. In FFPE mode, AmpliSeq generates 150bp amplicons. However, I believe these are tiled and it looks like there is less than 2X overlap. If I sequence using 2x75bp sequencing, would my targets be completely covered?
My concern is that there are "partially digested primers" between the adapters and the target sequence. If the amplicon is sequenced for 75bp from both ends, then that will leave a gap in the middle.
My concern is that there are "partially digested primers" between the adapters and the target sequence. If the amplicon is sequenced for 75bp from both ends, then that will leave a gap in the middle.
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