Qiime2 workflow

[kimshinseung (ss)]

Hi, qiime2 is a beginner.
I use paired-end seq data from illumina miseq.
Human intestinal microbial data is 16s ampliconsequencing.
I am curious about the workflow until OTU generation.

1. importing data ->qza

2. demultiplexing -> demux.qza

3. qualitycontrol
   r = 3, q ​​= 3, p = 0.75, n = o, c = 0.05 -> demux-filtered.qza
   We have progressed to this point.

4. At what stage does cutadapt go?

5. How does demultiplexing replace the completed fastq file with a qza file?

6. Is it correct to proceed with dada2 or deblur after proceeding to the present?

7. demux-filtered.qza. What is the command to move dada2 to a file?

8. Do you proceed with merge after denoise in dada2?

9. What if I try to use a database other than greengene data?

10. Do you generate otu via vsearch with the output of dada2? What is the command? 8. What is the format of the otu file?

I am wandering a lot with the first time.
Please give detailed explanation.
Thank you.

[colinbrislawn (Colin J Brislawn)]

Hello @kimshinseung,

Thanks for using Qiime 2! Welcome!

These are all really good questions, which is why they are covered in detail in the "Moving Pictures" tutorial. This tutorial shows a full qiime 2 pipeline, and you can compare this to the pipeline you described here. I think that once you have done the "Moving Pictures" tutorial, the order of steps will be more clear.
https://docs.qiime2.org/2019.1/tutorials/moving-pictures/

Qiime 2 makes use of new file types (.qza and .qzv) and I notice you have questions about those too. These are covered in detail here:
https://docs.qiime2.org/2019.1/tutorials/overview/

Let me know once you have finished the “Moving Pictures” tutorial or if you have any questions along the way.

Colin

[kimshinseung (ss)]

Thank you for answer.

1. Is the otu table in qiime1 the same as the feature table?
2. Is a database like greengene used in --m- below?
   ------------------------------------------------ qiime feature -table summarize
   –i-table table.qza
   –o-visualization table.qzv
   –m-sample-metadata-file sample-metadata.tsv
   qiime feature-table tabulate-seqs
   –i-data rep-seqs.qza
   –o-visualization rep-seqs.qzv
3. Does the trim-length of the deblur mean the minimum length of the faired-end seq to overlap the library size? Is this process a merge process?

Thank you

[colinbrislawn (Colin J Brislawn)]

@kimshinseung

Is the otu table in qiime1 the same as the feature table?

Yes. The features in the feature table could be OTUs or AVSs.

@kimshinseung

Is a database like greengene used in qiime feature -table summarize

Nope. That's for the --m-sample-metadata-file.
That's the first thing explained in the Moving Picture tutorial. Have you done the tutorial yet?

@kimshinseung

1. Does the trim-length of the deblur mean the minimum length of the faired-end seq to overlap the library size? Is this process a merge process?

From the Moving Picture Tutorial:

Next, the Deblur workflow is applied using the qiime deblur denoise-16S method. This method requires one parameter that is used in quality filtering, --p-trim-length n which truncates the sequences at position n .

I think so many of your questions will be answered when you do the tutorial! You will learn so much!

Colin