Prepare Metadata for pair-end sequencing data

[Huiyun_Wu]

Hi,

Can anyone give me an example to prepare metadata for pair-end sequencing data? The tutorial is for single-end sequencing data. https://docs.qiime2.org/2020.2/tutorials/moving-pictures/

Thanks!

[andrewsanchez]

Welcome to the forum, @Huiyun_Wu!

You can find examples of working with paired-end sequence data in a few different places.

Most importantly, keep in mind that you can get help directly from on the command line:

➜ qiime tools import --help
Usage: qiime tools import [OPTIONS]

  Import data to create a new QIIME 2 Artifact. See https://docs.qiime2.org/
  for usage examples and details on the file types and associated semantic
  types that can be imported.

Options:
  --type TEXT             The semantic type of the artifact that will be
                          created upon importing. Use --show-importable-types
                          to see what importable semantic types are available
                          in the current deployment.                [required]
  --input-path PATH       Path to file or directory that should be imported.
                                                                    [required]
  --output-path ARTIFACT  Path where output artifact should be written.
                                                                    [required]
  --input-format TEXT     The format of the data to be imported. If not
                          provided, data must be in the format expected by the
                          semantic type provided via --type.
  --show-importable-types Show the semantic types that can be supplied to
                          --type to import data into an artifact.
  --show-importable-formats
                          Show formats that can be supplied to --input-format
                          to import data into an artifact.
  --help                  Show this message and exit.
➜ qiime tools import --show-importable-types
Bowtie2Index
DeblurStats
DistanceMatrix
EMPPairedEndSequences

Note the output of the second command above was truncated for demo purposes.

The “Atacama soil microbiome” tutorial is a good next step after working through Moving Pictures
https://docs.qiime2.org/2020.2/tutorials/atacama-soils/

There is also a section in the metadata tutorial: Importing data — QIIME 2 2020.2.0 documentation

Let us know if you have any more questions

[Huiyun_Wu]

Thanks Andrew! I will try it out!

[thermokarst]

Something to keep in mind, @Huiyun_Wu is that there aren’t really any requirements in QIIME 2 regarding sample metadata (all of the old QIIME 1 requirements about linker-primer-sequence et al all gone):

https://docs.qiime2.org/2020.2/tutorials/metadata/

Most metadata-needs will be driven by your specific study. Sample metadata is the “secret sauce” that makes your sequencing data interesting, and specific to your study.

[Huiyun_Wu]

Hi Andrew,

I was following the “Atacama soil” tutorial https://docs.qiime2.org/2020.2/tutorials/atacama-soils/. The barcode fastq.gz file is already prepared and easy to download. However, I only have the forward and reserve fastq.gz file available for my project. I have the sample ID and barcode sequence information. Is there a tutorial illustrating how to prepare the barcode fastq.gz file?

Also, when I open the sample-metada.tsv for this tutorial, I can only find one column “barcode-sequence”, where is the other pair of barcode? Is it listed in a different sample ID? In another word, is BAQ1370.1.2, BAQ1370.1.3, BAQ1370.3 the same sample but with differrent barcode?

Thanks a lot!

[andrewsanchez]

@Huiyun_Wu

I only have the forward and reserve fastq.gz file available for my project. I have the sample ID and barcode sequence information. Is there a tutorial illustrating how to prepare the barcode fastq.gz file

It sounds like you might have multiplexed-paired end reads with the barcodes still in the reads, which means there is no need to create a barcodes.fastq.gz. However, without more information, we can't be sure. If the barcodes are in fact in your reads, you can use this tutorial as a guide:

Demultiplexing and Trimming Adapters from Reads with q2-cutadapt

NOTE This tutorial is a work in progress, and is incomplete at the moment. It demonstrates at a high level some of the methods available in the q2-cutadapt plugin available in QIIME 2 2018.2. Please stay tuned here for additional updates as this tutorial is expanded upon in the coming weeks. Multiplexed reads with the barcodes in the sequence reads can be demultiplexed in QIIME 2 using the q2-cutadapt plugin,…

@Huiyun_Wu

Also, when I open the sample-metada.tsv for this tutorial, I can only find one column “barcode-sequence”, where is the other pair of barcode? Is it listed in a different sample ID? In another word, is BAQ1370.1.2, BAQ1370.1.3, BAQ1370.3 the same sample but with differrent barcode?

I recently answered a question that spells out how the mapping between barcodes and sequences work here:

Moving pictures tutorial: Demultiplexing

Hi, @Rania, Welcome to the forum! Can you elaborate on what you mean by trying to allocate the barcodes inside the sequences? What exactly did you try? What were the expected results vs actual results? I think the observations noted below will help clarify some things: If you compare the sequence identifiers in both sequences.fastq and barcodes.fastq emp-single-end-sequences/sequences.fastq.gz and emp-single-end-sequences/barcodes.fastq.gz from the moving pictures tutorial), you…

Although the post is about single end sequences, the logic stands.