Demultiplex amplicon data on duel barcodes and priming sequence.

[JackieBadger]

Hi,

I am looking for a program that can de-multiplex ~1 million or more amplicon sequences based on 10bp barcodes at either end of the sequence. A set of barcodes are associated with the forward and reverse priming sequences, so these need to be taken into consideration in order to sort amplicons into the same orientation and allocate correctly.


It would also be massively beneficial if the program could quantify variants within and across samples.

Split_libraries in Qiime seems to only work on a single barcode (??)
This also goes to Fastx barcode splitter.

jMHC does exactly what I want but is only Java based and can not handle large Illumina sized data sets.

If anyone has any suggestions I would very much appreciate it.

[akbowser]

Hi JackieBadger,

I'm wondering if you ever found a solution to your problem?

I've never worked with NGS data before (still waiting for facility to sequence my stuff) but I also have barcodes associated with primers at both ends of my amplicons. Just trying to prepare myself for the long road ahead...

Thanks,

Kirsten

[JackieBadger]

Hey Kirsten,

Are you at Dal?
If so I can show you how to do it.
The short answer is that yes I managed to employ my old protocol using jMHC.
The issue was that the automatic memory allocation could not cope will huge data sets (compared to "moderate" 454 data sets). I tweaked the memory allocation and was able to process much larger data sets.

[akbowser]

Hi,
No, I'm at UNB (and unfortunately I don't know anyone here with NGS data experience).

I'm currently trying to teach myself Qiime, as my ultimate goal is to assign OTUs to my 454 sequenced amplicons. I don't know too much about Qiime yet, but based on your post it seems it won't allow me to demultiplex dual barcodes. Any suggestions on how I should go about dealing with my data? (keeping in mind that I am a complete novice and will be learning everything from scratch)
Thanks for your input,

Kirsten

[JackieBadger]

send me an email and I'll give you some pointers [email protected]