Hello Gisella,

maybe you should read this one: http://ddgehelp.blogspot.de/2005/11/dgge-guide.html

I think it depends on the length of your PCR product. Some sequencing reactions cannot be done for the whole Fragment, so it would be favourable to also have the possibility of sequencing the Fragment from the other site. Both sequencing reactions together can give you the complete sequence of your Fragment.

I agree with Daniela. In my experience, I find that when my fragment to be analysed is 700bps then, if using only one primer, the sequencing reaction starts to become very inefficient close to the end of the sequence so in this case its better to use 2 primers. If its a really big fragment then you may need more than one set of primers and use a shotgun approach. It also depends on the type of sequencer used as I believe the next gen sequencers are much more efficient. Hope that helps

Your question is very confusing. In general, if you do a good clean-up of the PCR products prior to sequencing, you can use the same primers for sequencing that you used for the PCR (of course, you will only use one primer in a sequencing reaction to obtain the sequence of one strand.) Whether you should sequence both strands of your products depends on the read length that you need and the quality of the traces in the regions where critical variations occur. It's also important to keep in mind that primers are cheap. So if you want to use primers for sequencing that don't have the GC clamp of the primers used for PCR/DGGE, you can easily order some (from IDT for example).

https://www.idtdna.com/pages/products/dna-rna/custom-dna-oligos

If you mean by the two sets of primers as the outward and inward primers then...Simply.....you will be sequencing in both directions and this is your guaranty for the accuracy of your reading of the base pairs.................One strand is not enough! and more over it helps to cover the end bit of the sequenced fragment specially if you have a very long fragment .