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I am interested in modifying my standard PE adapters with indexes during the final amplification step. Has anyone tried this? Have you heard/read anything like this before?
It seems the Read2 sites are different in the 'standard' vs 'multiplexed' adaptors (whereas the Read1 sites are unchanged).
I was wondering if there is some reason why a reverse-complement of the Read2 site in the 'standard' adapter would not serve as a suitable Index Seq Primer.
For example, I could introduce an index into my libraries by amplifying standard libraries with a reverse primer such as "5' P7_Site--Index--Standard_Read2_Site." Then use a 'custom' primer for the Index Seq Primer (that is the reverse complement of the standard Read2 site).
However, I am wondering why the Read2 site is different in "standard" versus "multiplexed" adapters. Perhaps Illumina changed the Read2 site for the multiplexed adapters because it does not serve as a good Index Seq Primer?
Any ideas would be greatly appreciated. Thanks in advance!
It seems the Read2 sites are different in the 'standard' vs 'multiplexed' adaptors (whereas the Read1 sites are unchanged).
I was wondering if there is some reason why a reverse-complement of the Read2 site in the 'standard' adapter would not serve as a suitable Index Seq Primer.
For example, I could introduce an index into my libraries by amplifying standard libraries with a reverse primer such as "5' P7_Site--Index--Standard_Read2_Site." Then use a 'custom' primer for the Index Seq Primer (that is the reverse complement of the standard Read2 site).
However, I am wondering why the Read2 site is different in "standard" versus "multiplexed" adapters. Perhaps Illumina changed the Read2 site for the multiplexed adapters because it does not serve as a good Index Seq Primer?
Any ideas would be greatly appreciated. Thanks in advance!