I am trying to understand the theoretical bases and practical causes for why R2 is poorer quality compared to R1 for Illumina Paired End sequencing.
While there are some posts here that discuss this observation: MiSeq: R1 vs. R2 reads show big quality difference - is that normal? Should I trim both R1 and R2 if only R2's quality is poor?
what I am most interested is in understanding WHY this happens. Could someone point me in the right direction please - specific papers, or an Illumina technical note, a Youtube presentation, etc. - anything useful. THANK YOU!
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written
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Anand Rao
2 answers
The Illumina paired end sequencing involves a temporal separation and a mechanistic process.
The "first in pairs" are sequenced first then the fragments "bend" over, reattach, then reads in the second pair are sequenced on the same flow cell surface.
Depending on the protocol this second step may occur days later. Hence it is not unexpected to have lower qualities on read 2.
See this thread on SA and my answer there.
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