universal 18s eukaryotic primers

[giderk]

I'm wondering if anyone has identified a good primer pair that spans the 18s rRNA v4 region and is proven to work well on the MiSeq platform. I haven't found any established protocols in the literature yet. Most of the recent publications from 2014 refer to 454 sequencing. I'd be interested in other primer suggestions spanning v5, v7 and v8 as well. I've found this paper useful, "Characterization of the 18S rRNA Gene for Designing Universal Eukaryote Specific Primers" by Hadziavdic et al., but again, all these primers were designed with 454 in mind.

If anyone is willing to share universal eukaryotic 18s primers and amplification protocols they have found to work well on the MiSeq, I'd be really appreciative.

[cecay]

Don't know if this helps, but take a look here:

http://www.earthmicrobiome.org/emp-standard-protocols/18s/

[thermophile]

The biggest concern for 18S (even more so than 16S) is to get complete overlap of forward and reverse since most people looking at Euk SSU are wanting to talk about single base changes. Without complete overlap, your quality won't be high enough to consider single bases. So look at the available primers and choose a few sets that seem to be roughly 250-300bp long (miseq v2 500 cycle or v3 600 cycle). Do some in silico pcr against a big database like Silva and see if the "universal" primers are universal enough for your groups or if they are biased against things that you are interested in.