Friday 26 October 2018

Spiking sequencing libraries with PhiX

If you are sequencing a low complexity library (e.g. a PCR-amplicon library) using Illumina sequencers, it's a good idea to spike in some PhiX to improve base quality, as explained here and here on the Illumina support pages. On the seqanswers forum they say that PhiX is used for:
"introduction of sequencing diversity in low-complexity libraries (diversity is needed to discriminate clusters and create signal thresholds for base-calling). As the software has improved, the recommended amount of phiX spike-in has decreased." 
Another comment says:
"Older basecalling software on the MiSeq used to require a 50% phiX spike for low-diversity (amplicon samples). More recently, the software has been updated and only requires a 5-10% spike."

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