I am new to microbial analysis and got stuck with my analysis. I have install qiime2-2022.2 in my conda env. I have RNAseq data obtained using shoreline Biome complete kit (strain ID) which is a pacbio product. I followed their procedure to do advance ASVs analysis which uses DADA2 pipeline.R in terminal. The ASV were classified using Athena database provided in SBanalyzer software. The files generated were then imported into qiime2 to do further analysis by using following code:
-Import the ASV .tax feature taxonomy into Qiime2
qiime tools import --input-path <path to .tax file> --type 'FeatureData[Taxonomy]' --input-format HeaderlessTSVTaxonomyFormat --output-path taxonomy.qza
<path to .tax file> is the absolute path to the ASV .tax file generated by sbsearch.exe
-Import the .biom feature table into Qiime2
qiime tools import --input-path <path to .biom table> --type 'FeatureTable[Frequency]' --input-format BIOMV100Format --output-path table.qza
<path to .biom table> is the absolute path to the .biom ASV frequency table file generated by DADA2
-Import the ASV .fasta file into Qiime2
qiime tools import --input-path <path to ASV .fasta> --type 'FeatureData[Sequence]' --output-path rep_seqs.qza
is the absolute path to the ASV .fasta file generated by DADA2
After this I followed moving picture tutorial.
I made feature table using above files. I followed moving picture tutorial for generating tree for phylogenetic diversity and alpha and beta diversity . The question that I have is:
the observed ASV in my sample is too low (eg: lowest to 8 ASVs)
I directly plotted taxonomy barplot using taxonomy files that was imported to qiime2 (taxonomy.qza). I am not sure if I need to make taxonomy classifier. As the ASV were classified using athena database, can I use SILVA trained classifier? How do we make 99% or 97% ASV sequence and 99% or 97% ASV taxonomy?
I read through training feature classifiers with q2-feature-classifier but did not see anywhere that mention the percentage.
how do I get how many ASVs reads were there in each sample?