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lenaarenot committed Jun 19, 2024
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Microbiome analysis using amplicon sequencing is central to many ecological studies.
The produced amplicon sequencing data are converted to OTU tables and represent the input
for the ampvis2 tool, where it can be visualised in various ways.`{% cite Andersen2018 %}`
for the ampvis2 tool, where it can be visualised in various ways.{% cite Andersen2018 %}
If you already have amplicon data produced and ready to feed in and visualise it,
then you can start with this tutorial. First of all you can put your data into a
rarefraction curve to explore reads against number of OTUs. Than you can input your
data into subsets and finaly create a heatmap, or a boxplot, or an ordination plot
or even a timeseries plot out of it. Most of them are described in
`{% cite ampvis-intro %}`
{% cite ampvis-intro %}
![overview of visualisation methods](./images/overview.png
"Overview of posible visualisation methods (taken from: Introduction to ampvis2 by Kasper Skytte Andersen)")
Your data need to be in an acceptable format for the ampvis_load tool. The tool
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## heatmaps
We now can use our data, put them in subsets and create ungrouped, or grouped output or
even grouped with facets.
The subsets are based on variable we set and available in the metadata. `{% cite Andersen2018 %}`
The subsets are based on variable we set and available in the metadata. {% cite Andersen2018 %}
Note: in the next sections, we give you prepared workflows on Galaxy and the set of parameters to choose
for running the indicated workflow. But some parameters are pre-chosen for you e.g. taxonomic level to
aggregate the OTUs.
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>
{: .hands_on}
![Running the workflow](./images/heatmap_no_group.png "Running the workflow,
choose the right datasets and mandatory parameters")
![Running the workflow](./images/choose_parameters.png "Running the workflow, choose the right datasets and mandatory parameters")
Choose the metadata variable as Plant and metadata values as Aalborg East & Aalborg West.
In the next box you can see the resulting heatmap.
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>
> Result of the heatmap created with ungrouped data.
>
>![Result of the heatmap](./images/choose_parameters.png "Result of the heatmap")
>![Result of the heatmap](./images/heatmap_no_group.png "Result of the heatmap")
{: .details}
> <details-title> metadata values error while running workflow </details-title>
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### heatmap (grouped)
You can find the workflow "ampvis2 heatmap v2.0 (only group)" on Galaxy and use it for the tutorial.
We used 2 different metadata subsets:
1) metadata we used for this subset: metadata variable = Plant and metadata values = Aalborg East & Aalborg West.
1) metadata we used for this subset: metadata variable = Plant and metadata values = Aalborg East & Aalborg West
plus grouped by = Plant
2) metadata we used for this subset: metadata variable = Period and metadata values = Winter & Summer.
2) metadata we used for this subset: metadata variable = Period and metadata values = Winter & Summer
plus grouped by = Year
> <hands-on-title> Run a workflow </hands-on-title>
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### heatmap (grouped with facets)
You can find the workflow "ampvis2 heatmap v1.0 (group+facet)" on Galaxy and use it for the tutorial.
We used 2 different metadata subsets:
1) metadata we used for this subset: metadata variable = Plant and metadata values = Aalborg East & Aalborg West.
1) metadata we used for this subset: metadata variable = Plant and metadata values = Aalborg East & Aalborg West
plus grouped by = Plant and facet by Period
2) metadata we used for this subset: metadata variable = Period and metadata values = Winter & Summer.
2) metadata we used for this subset: metadata variable = Period and metadata values = Winter & Summer
plus grouped by = Year and facet by Period
Expand All @@ -213,19 +212,17 @@ We used 2 different metadata subsets:
>
> Result of the first metadata subset heatmap created with grouped by Plant data and facet by Period.
>
>![Result of the heatmap](./images/heatmap_plant_period.png "Result of the heatmap created with
> grouped by _Plant_ and facet by _Period_")
>![Result of the heatmap](./images/heatmap_plant_period.png "Result of the heatmap created with grouped by _Plant_ and facet by _Period_")
>
> Result of the second metadata subset heatmap created with grouped by Year data.
>
>![Result of the heatmap](./images/heatmap_year_period.png "Result of the heatmap created with
> grouped by _Year_ and facet by _Period_")
>![Result of the heatmap](./images/heatmap_year_period.png "Result of the heatmap created with grouped by _Year_ and facet by _Period_")
>
{: .details}
## ordination plots
We now can use our data, put them in subsets and create different plots by using different ordination methods.
Like with heatmaps, the subsets are based on variable we set and available in the metadata.`{% cite Andersen2018 %}`
Like with heatmaps, the subsets are based on variable we set and available in the metadata.{% cite Andersen2018 %}
Note: in the next sections, we give you prepared workflows on Galaxy and the set of parameters to choose
for running the indicated workflow. But some parameters are pre-chosen for you e.g. ordination method,
transformation (if used) and others like to colour and label the points or frames.
Expand Down Expand Up @@ -280,7 +277,7 @@ Metadata we used for this subset: metadata variable = Plant and metadata values
## boxplot
We now can use our data, put them in subsets and create a boxplot.
Like with heatmaps, the subsets are based on variable we set and available in the metadata. `{% cite Andersen2018 %}`
Like with heatmaps, the subsets are based on variable we set and available in the metadata. {% cite Andersen2018 %}
Note: in the prepared workflow on Galaxy we provide in this tutorial some parameters are pre-chosen for you
e.g. number of taxa to show. The samples are grouped by _Period_.
Metadata we used for this subset: metadata variable = Plant and metadata values = Aalborg East & Aalborg West.
Expand Down

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