I am doing RNAseq and would like to pool my library with some other lab members. The problem is we are not using the same library preparation kit (and thus the index). I would like to know whether there are any incompatible concerns?

I check the index for these kits, I would like to know what should I look for?
Here is my checklist:

  1. check whether there are any identical indexes (which I use the index given in both user manual)
  2. check whether the indexes are identical after trimming off 1 base from either end

My question is do I also need to check the complement version of these indexes? And also the reverse and reverse complement?

One of my kits provides a unique Dual-indexed adapter: P7 index sequence, P5 index sequences (1 and 2) ~~ this is from the KAPA kit. On the other hand, my lab member is using the NuGEN Ovation® SoLo RNAseq kit, which only provides one index sequence (of each well) for me.

May I know which sequence I should make the comparison to?

The final question is I found that one of the "reversed" indexes from the SoLo kit is identical to one of the indexes I used after trimming one 5' base.

SoLo original index: ACCATCCT
SoLo reversed index: TCCTACCA
SoLo reversed and trimmed: CCTACCA  #overlap found with this to next index sequence
KAPA P7 Index sequence: CCTACCAT

I know that the SoLo kit claim they will have 8 random bases following the provided index. Do you think if there is a random T following the CCTACCA will cause a problem? Or the demultiplexing algorithm can deal with the issue by knowing that the index is actually TCCTACCA T rather than T CCTACCAT. Of course, this is not a problem if I do not need to compare the index with its reverse sequencing in the counterpart.



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