Using illumina transposon creation to get by amplicon length limit?

mothur bugs
1

Hi all;

I don’t really know where to put this post. I guess bugs is as good as any, as this references somewhat to the longer, non-overlapping reads issues on the 300bp 2x miseq where diversity gets artificially amplified at sequence ends.

I was wondering if it was possible to use nextera’s transposon kit to break down larger amplicons into smaller sizes that can be then reassembled (as used in typical shotgun metagenomics)? I’ve read a few papers in which this general approach seemed to work.

I only ask because [relatively] universal archaeal primers are hard to come by, and the best ones I’ve seen create pretty big (~700 bp) amplicons.

Any thoughts?

2

Hi,

At this point, mothur is not set up to work with randomly sequenced regions of the 16S rRNA gene.

Pat

3

Sorry I should’ve been more specific:

using another program for assembly, could the resulting sequences be used for analysis in mothur? In theory, this should get rid of the inflated diversity from unpaired read ends, reducing the memory required for the distance matrix and clustering steps (or am I missing something here)?

Additionally, how drastically might this assembly affect the resulting community structure as opposed to no-assembly-required, single-read 16S data?

Wish there were more consensus high throughput sequencing parameters for archaea.

thanks

Jon