Understanding the primer design output

I have run primer design, it ran successfully, and now I am trying to understand the output. I am specifically looking at the *.primer.summary file. Here’s what I see in my file :

  • The first line in the file identifies the OTU of interest and lists all the members.

The second line then lists a header, which is_primers, mintm, max tm_. On the subsequent lines it proceeds to list a few primers.

Once that’s done there is new table, which has the following header:OtuNumber, Primer, Start, End, length, mismatches, minTm and maxTm.

I understand Start, End, length, mismatches as well as minimum and maximum melting temperatures. What I don’t understand are the way the primers are outputted and why there are two separate lists of primers that contain different information.

My question about the primer : In the first list of primers, which only lists the melting temperatures, I assume that those primers are specific to my OTU of interest, but is there some order to them? Specifically, does this method output forward & reverse primers, and how are they represented? Am I missing information in the output?

For that second list below : what is it’s purpose and what is it trying to tell me? Are those primers for other OTUs? Is there a relationships between the primers of my OTU of interest and other OTUs and this table is trying to tell me something about it?

Thanks for any help that you offer and I apologize for ignorance.

I eventually decided to look at the source code for primer design, and this is what I’ve gotten from the code:

The first table is just the primers, and I really didn’t see any indication of what is forward or reverse, specifically. So I am still not certain on this point.

The second table is : “check each otu’s conseq for each primer in otunumber” I think this is measure of OTU specificity. I think in a perfect world, I would like that table to be very very small. I’ve also noted that at least one of the suggested primers doesn’t seem to appear too often in that second table (meaning it isn’t found in other OTU sequences), so I take this to be a good thing, and that I probably should use that.

I tested the pairs in the silva test primer module. It didn’t seem to be picky about which was forward and which was reverse, it was fairly specific for the organism.