Hi everyone, I am very new to QIIIME 2 (with no prior experience in microbiome bioinformatics) and I would like to seek advice on how to correctly import paired-end reads as shown below.
I have gone through the attacama and import tutorials.
I tried running the manifest format only to realize that it only works for demultiplexed reads as the error output was:
There was a problem importing /Users/ega/Augmentin/pe-64-manifest:
/Users/ega/Augmentin/pe-64-manifest is not a(n) PairedEndFastqManifestPhred64V2 file:
Does this also validate that my reads are multiplexed? (pardon me if this sounds like a redundant question)
I also tried the EMP multiplexed paired-end protocol, of which requires 3 files, namely, forward, reverse and barcode sequences. And I am unsure how to ‘merge’ all the forward reads from different samples into a single file as the tutorial comes with ready-to-import forward, reverse and barcode sequences.
The sequencing files are in the respective folders shown here:
These are the files shown in S1D0:
Barcode sequence is also provided by the sequencing company in a .xls file:
These are the notes provided by the sequencing company:
Hence, from the above notes, _1.fq.gz and _2.fq.gz files seems to have their barcodes and primer sequences removed.
Do i have to ‘merge’ all the forward and reverse reads from different samples into a single forward and reverse file ? (like how the attacama tutorial does it)
How do i create a barcodes.fast.qz from the .xls file (if it is even required at ll)
If so how do i do it. If not, what are some alternatives to import the sequences given my specific circumstances.
Will appreciate all advice and feel free to ask clarifying questions for a better understanding of my current situation.