Hi,could you please help me out,thank you !!
I am using qiime2 2024.2,and need to analyze 16s(archaea), using the protocol,and specific primer, and I can not get any archaea left. Here are the codes.
Can you please provide more details. Are you not seeing any archaea at all, or just fewer than you expect?
What samples are you sequencing from?
Note: You do not need to follow all the steps from that tutorial, as it is just a set of examples, as described there. You can try skipping the rescript filter-seqs-length-by-taxon step. See this example and this example. I say this as that step might be removing sequences that you need for classification. Or, worst case scenario there are few archaea in your sample.
Have you tried GTDB or RDP? You can download both via RESCRIPt.
Thank you
I am not seeing any archaea at all
My sequencing are 16S rRNA sequencing sequencing,the RNA was extracted from human stool, and used the primer"519f-915r".
I am sure the sample have archaea
Thank you
Here is code I used for dada2
echo -e 'sample-id\tforward-absolute-filepath\treverse-absolute-filepath' >/data/gn/file01/manifest.tsv
ls R1.fq.gz|while read id;
do
echo "${id%%},$PWD/$id,PWD/{id%%_*}_R2.fq.gz" >>/data/file01/manifest.tsv;
done
sed 's/,/\t/g' /data/file01/manifest.tsv > /data/file01/manifest_pe.tsv
How good are your quality plots? I ask because it is quite rare to use --p-trunc-len-f 0 --p-trunc-len-r 0 unless the quality is quite high. I'd recommend specifying other truncation values. Can you share your dada2 stats QZV file?
That looks really good. Perhaps try playing with the --p-min-fold-parent-over-abundance parameter, to increase your "non-chimeric" read count. This is discussed here.