Dear All, I have a question. I have got paired reads from the company. The sequence base for each forward and reverse read truncated at 300 each. Then I use Greengenes (16S rRNA) 13.8 marker gene database using following commands. qiime feature-classifier extract-reads --i-sequences 99_otus.qza …

Hi @Aqleem12 ! Taxonomic classification of features is largely the same for single-end vs. paired-end reads. You'll first need to denoise your paired-end sequences with DADA2, which will join your reads for you and produce a feature table and representative sequences. The Atacama soil microbiome tuto…

Dear Sir, Thank you for your reply. I did denoise as follow with DADA2, qiime dada2 denoise-paired --i-demultiplexed-seqs demux.qza --o-table table --o-representative-sequences rep-seqs --p-trim-left-f 0 --p-trim-left-r 0 --p-trunc-len-f 300 --p-trunc-len-r 300 Now I want to train c…

[image] Aqleem12: –p-trunc-len 600 Please see my previous reply about trunc-len: [image] jairideout: When classifying representative sequences that came from joined paired-end data, it’s recommended to not supply a truncation length (so in your command, you can remove --p-trunc-len 300).…

Dear Sir, Thanks for your reply. I use the primers with 85% Greengene reference sequences. They work well and quickly. The genus and species were absent in that 85%Greengene . However when I run the primers with 99%Greengenes it took almost two days, however disconnected and could not give me the r…

[image] Aqleem12: Thanks for your reply. I use the primers with 85% Greengene reference sequences. They work well and quickly. The genus and species were absent in that 85%Greengene . That's great news, thanks for trying that out! [image] Aqleem12: However when I run the primers with 99%…

I split off @Aqleem12 's latest reply into a new forum topic because there's a question in there that is better suited for its own topic. Following up here to confirm that @Aqleem12 was able to successfully extract reads from 99% Greengenes reference sequences, it took around 12 hours to complete.