Analysis with QIIME

Description

Before we begin the analysis of the PICRUSt predicted metagenome, we must first change a few parameters, so that QIIME can recognize the .biom file as a valid format. See Section 3.1 for more information.

For PICRUSt analysis, we are going to generate the parameters file in Terminal. The command below will generate a new file in the current working directory called picrust_parameters.txt. If you want a different level, you can change the last line accordingly.

Download PICRUSt parameters file here or run the command below

echo 'summarize_taxa:md_identifier "KEGG_Pathways"' >> picrust_parameters.txt
echo 'summarize_taxa:absolute_abundance True' >> picrust_parameters.txt
echo 'summarize_taxa:level 3' >> picrust_parameters.txt
echo 'beta_diversity:metrics bray_curtis,euclidean' >> picrust_parameters.txt

A. Summary Plots

Next we will run plot the summary of KEGG levels using QIIME commands. We will need to reference the parameters file we just created in the previous step. If you are using Level 3 for the PICRUSt your parameters file must state summarize_taxa:level 3 but if you are using Level 2, it must state summarize_taxa:level 2

summarize_taxa_through_plots.py \
-i predicted_metagenomes_L3.biom \
-o sum_taxa_level3/ \
-p picrust_parameters.txt

B. Beta Diversity PCoA

Step 1: Get summary of counts

biom summarize-table \
-i metagenome_predictions.biom \
-o metagenome_predictions_stats.txt

Step 2: Rarefy data Using the depths found in the previous commands output, rarify the PICRUSt table to the minimum sequencing depth across all samples.

single_rarefaction.py \
-i metagenome_predictions.biom \
-o metagenome_predictions_even_sampled.biom \
-d MINIMUM

Step 3: Run beta diversity analysis and visualize with PCoA.

beta_diversity_through_plots.py \
-i metagenome_predictions_even_sampled.biom \
-o bdiv_metagenome_predictions/ \
-m mapping_file.txt \
-p picrust_parameters.txt