Hello.
I use qiime2 2020.6. I have dataset with about 410 samples. In DADA2 step, when truncating 150 bp, I loose many samples and just 70 samples remain because many of reads have 126 bp length.when I truncate 126 bp I have all samples but in taxonomy step (gg-13-8-99-...) the result is only unassigned and k-bacteria(without any taxa levels) .I was wondering why it happens and what I should do.
Hi @Fatemeh_Abedini,
Could you give us some more details about your hypervariable region and target gene and the commands you ran? Where did you get the classifier?
Best,
Justine
Hi
Five regions of the 16S rRNA gene.
primers:F1-TGGCGAACGGGTGAGTAA
F2-ACTCCTACGGGAGGCAGC
F3-GTGTAGCGGTGRAATGCG
F4-GGAGCATGTGGWTTAATTCGA
F5-GGAGGAAGGTGGGGATGAC
R1-CCGTGTCTCAGTCCCARTG
R2-GTATTACCGCGGCTGCTG
R3-CCCGTCAATTCMTTTGAGTT
R4-CGTTGCGGGACTTAACCC
R5-AAGGCCCGGGAACGTATT
I used "gg-13-8-99-515-806-nb-classifier" in qiime2.(also I try with gg-13-8-99-nb-classifier but same result)
DADA2 command:
qiime dada2 denoise-paired
--i-demultiplexed-seqs paired-end-demux.qza
--p-trim-left-f 18
--p-trim-left-r 18
--p-trunc-len-f 126
--p-trunc-len-r 126
--o-table table.qza
--o-representative-sequences rep-seqs.qza
--o-denoising-stats denoising-stats.qza
when I truncate 150bp ,70 sample from 476 remain but the taxonomy result is good
.
Hi @Fatemeh_Abedini,
You need to use the full length classifier because those are not the 515-806 region and so the classifier wont work as well. Multiple region scaffolding is a hard problem. Are those the SMURF primers? If I were you, I my recommendation today would be to look at the ion torent thread:
Best,
Justine
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