Background:
Instead of combining 4 50 cycle kits for our HiSeq dual index runs, we typically use standard 200 cycle kits and 1 50 cycle kit to top off up to 4 dual index runs starting on the same day. Further, we split one incorporation mix between FCA and FCB at the start of all our HiSeq runs, and split the second mix at the turn.
Our problem:
On this particular dual index run, the reagents were not topped off at the start of the run, thus the incorporation mix ran dry sometime during the dark cycles, causing index2 to fail. The dilemma here is that simply adding incorporation mix and resuming the run at the beginning of index2 will cause the read to be out of phase since an unknown number of bases were added during the dark cycles before the mix ran dry. Since the index2 primer is grafted to the flowcell we cannot denature the strand and start the read over either.
Any suggestions as to how to save this run? Thanks in advance!
Instead of combining 4 50 cycle kits for our HiSeq dual index runs, we typically use standard 200 cycle kits and 1 50 cycle kit to top off up to 4 dual index runs starting on the same day. Further, we split one incorporation mix between FCA and FCB at the start of all our HiSeq runs, and split the second mix at the turn.
Our problem:
On this particular dual index run, the reagents were not topped off at the start of the run, thus the incorporation mix ran dry sometime during the dark cycles, causing index2 to fail. The dilemma here is that simply adding incorporation mix and resuming the run at the beginning of index2 will cause the read to be out of phase since an unknown number of bases were added during the dark cycles before the mix ran dry. Since the index2 primer is grafted to the flowcell we cannot denature the strand and start the read over either.
Any suggestions as to how to save this run? Thanks in advance!
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