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Old 05-09-2019, 04:35 AM   #2
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Location: USA, Midwest

Join Date: May 2008
Posts: 1,177

It sounds like your new lab is following the protocol developed by Patrick Schloss' lab (or something very similar) for construction dual indexed Illumina amplicon libraries. You can review the full details of the protocol here. Our core follows this protocol for 16S-V4 amplicon library prep and it's very reliable.

Regarding the Pad and Linker, these are described in Appendix C (Primer Design) of that document:

The pad is a 10-nt sequence to boost the sequencing primer melting temperatures. The link is a 2-nt sequence that is anti-complementary to the known sequences.
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