Orienting Reads with classifier-consensus-vsearch

I am analyzing Illumina 16S data in QIIME 2. I have created a script by combining the Atacama Soil Tutorial and the Moving Pictures Tutorial. After creating a barplot from my reads I see that 50% of the sequences are assugned "kingdom bacteria"

I have figured out that this is because my reads are not oriented in the same direction. I have read a couple forums and from what I have seen, the issue is that classify-sklearn command requires the reads to be in the same orientation. I think I need to use the classifier-consensus-vsearch command instead. In my code i am currently using the gg-13-8-99-515-806-nb-classifier.qza from the Moving pictures tutorial. Here is the code that I run in the sklearn command

qiime feature-classifier classify-sklearn
--i-classifier gg-13-8-99-515-806-nb-classifier.qza
--i-reads rep-seqs-short.qza
--o-classification taxonomy.qza

In order to orient all the reads the same way I need to add reference reads and reference taxonomy. What is the easiest way to tweak this code to orient my reads the same way? The classifier included in the code above is a reference taxonomy, correct? Do I need to create my own reference reads? Please let me know if I need to clarify any of my questions. Thank you for taking the time to read and respond, I really appreciate it!

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Hi @Nicolette_HernandezK,

If you search the forum you'll find several posts, like the ones below about reorienting reads using RESCRIPt:



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