Primer sequences in an oligos file

Theory behind mothur
1

In creating an oligos file, why is forward primer treated as reverse and vice versa?

In text, “The forward primer is best thought of as the forward sequencing primer. So if you are using the 16S rRNA primers 27f and 338r to generate sequencing substrate, but you are sequencing off of the 338r end of the fragment, you would list 338r as the forward primer and 27f as the reverse.” is wrtten, but it does not make sense.

Does anybody help me out?

2

That’s because the sequencing is done in the opposite direction of what we typically think of doing in 16S. So the first primer you’ll see is the reverse PCR primer. But because it’s attached to the forward sequencing primer, we think of it as the “forward” primer. Clear as mud? We don’t use the “reverse” primer option.

Pat