some questions about the ITS data from MiSeq run

Question

[hua.peng1314]

I have a 2*300 Miseq run which produced two fastq files;The data is from ITS region with the primer ITS1 and ITS4.The Mothur and Qiime software are needed to combine the two fastq files depend on their overlap. I know the length of the whole ITS region have hight-variability.

It's inevitable that some ITS regions are longer than 600bp.I am not sure how to deal with them.

Any software can combine them for the followed analysis with Mothur or Qiime.I tried also makecontigs in mothur. But mothur joined ALL reads, so I was suspitios about the ouput.I also use some other software and approx half of the reads joined.It means I will lose half of the information.More serious I can't get any information about the species whose ITS region is longer than 600bp.Any suggestion? Thanks.

Answer 1

[marina.v.yurieva]

Can you just analyse ITS1 and ITS4 independently?

You can also try to join them using something like flash to see how well do they join (if it's more than 600 bp they shouldn't join at all!).