cutadapt / trim-paired / option “front” and “adapter” – User Support

Hello,

I have sequences in paired end that have both primers in R1 and R2 files.
For exemple, a sequence in the forward ( R1 ) file :
In bold the 2 primers :

GTACACACCGCCCGTCGCTCCTACCGATACCGGGTGATCCGGTGAACCTTTTGGACCGTTTTTCGGAAAAATAAGTAAACCATATCACCTAGAGGAAGGAGAAGTCGTAACAAGGTTTCCGTAGGTGAACCTGCAGAAGGATCAAGACCAAGTCTCTGCTACCGTACGTCTTCTTAATCTCGTATGCCGTCTTCTGCTTGAAAATTGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG

A sequence In the reverse ( R2 ) file:

TGATCCTTCTGCAGGTTCACCTACGGAAACCTTGTTACGACTTCTCCTTCCTCTAGGTGATATGGTTTACTTATTTTTCCGAAAAACGGTCCAAAAGGTTCACCGGATCACCCGGTATCGGTAGGAGCGACGGGCGGTGTGTACTGTAGAACCATGTCGTCAGTGTAGATCTCGGTGGTCGCCGTATCATTAAAAAACGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG

I wish to remove the primers to keep the sequence inside so I used cutadapt like that:

qiime cutadapt trim-paired 
--i-demultiplexed-sequences demux-paired-end.qza 
--p-front-f GTACACACCGCCCGTC  
--p-adapter-f GTAGGTGAACCTGCAGAAGGATCA  
--p-front-r TGATCCTTCTGCAGGTTCACCTAC 
--p-adapter-r GACGGGCGGTGTGTAC 
 --p-discard-untrimmed 
--verbose 
--o-trimmed-sequences trimmed_remove_primers.qza 

After that I can see my trimming sequences with this command:

 qiime tools extract /
--output-path trimmed_remove_primers /
--input-path trimmed_remove_primers.qza

In the forward ( R1) file, unfortunately I have always the forward primer but the reverse primer (and the downstream sequence) has been correctly removed:

GTACACACCGCCCGTCGCTCCTACCGATACCGGGTGATCCGGTGAACCTTTTGGACCGTTTTTCGGAAAAATAAGTAAACCATATCACCTAGAGGAAGGAGAAGTCGTAACAAGGTTTCC

In the reverse ( R2 ) file, the reverse primer has been correctly removed but the forward primer with the downstream sequence is always here:

GTACACACCGCCCGTCGCTCCTACCGATACCGGGTGATCCGGTGAACCTTTTGGACCGTTTTTCGGAAAAATAAGTAAACCATATCACCTAGAGGAAGGAGAAGTCGTAACAAGGTTTCCGTAGGTGAACCTGCAGAAGGATCAAGACCAAGTCTCTGCTACCGTACGTCTTCTTAATCTCGTATGCCGTCTTCTGCTTGAAAATTGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG

I don’t undersand why only the reverse primer are removed on the forward (R1) and reverse (R2) file?

After that removing primers step I will use dada2, this is correct right?

Thank you for any advice and have a nice day :slight_smile:
Jérémy Tournayre

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