I'm currently using pyRAD to conduct some population genetic analysis on my sequence data. I have paired-end sequence data of RAD-seq by Illumina Hiseq X and their adapter sequences had already been trimmed off when they arrived. (So I don't have the barcode data of them.) So I skipped step 1 as manual says and went straight to step2, but i got these lines below.
pyRAD -p params.txt -s2
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pyRAD : RADseq for phylogenetics & introgression analyses
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step 2: editing raw reads
No demultiplexed files found. Check path.
I'm wondering why the program shows this. Of course, I have checked the path to my fastq files. I can't get what is wrong with my params.txt or the input files only by this short error message.
I have additionally 2 questions here.
1)What's the difference between "Demultiplex" and "sorted"? Sometimes these words are used in very similar, but not in the same ways, I guess. My data is already trimmed off(=Demultiplexed), but the manual says I can't skip step1 unless my data is sorted. So, does this error means I can't skip step1 because my data isn't sorted yet? If so, how can I fix it?
2)How do you usually make sure your paired-end data is demultiplexed or not?
I'm really new to this program and I can't find any similar problems to mine on the web. Any advice would be appreciated. Thanks.