Greg Bonito kindly prepared a Qiime cheat sheet that you can use in addition to the Qiime tutorial that’s on line….
Microbial Ecology & Evolution 2/16/12
Analyzing 454 Data with Macqiime
Before we get started, there are some things you should know…
QIIME and other bioinformatic programs work with through Unix command line interface. This may sound intimidating, but if you take a few hours to learn some basic operations in your Unix/Linux/Terminal, it will save you *LOTS* of time and frustration in the future. There are plenty of great free online tutorials to help you out. Here’s one the first of ~6,000,000 google hits for “unix tutorial”: http://www.ee.surrey.ac.uk/Teaching/Unix/
Also, you will want to have a good text editor to view, edit, and check your files. TextWrangler is a good free editor: http://www.barebones.com/products/textwrangler/. If you have any troubles with input files with Qiime, check to make sure that files are saved with Unix (LF) Line breaks and Encoding with Unicode (UTF-8), and that there are no hidden endings on your file.
Getting Started
Open your Terminal (find in your Utilities folder in your Applications). In your terminal, navigate to your “qiime_tutorial-v1.4.0” folder. On my machine I type:
gregorybonito$ cd /Users/gregorybonito/macqiime/qiime_tutorial-v1.4.0
(My computer is named “gregorybonito” and the “$” indicates the UNIX prompt, so I don’t actually type those. The UNIX command “cd” stands for “change directory”. You can save yourself the trouble of typing the entire path by locating the file in your “finder” or windows directory and dragging it into your terminal. The path, as if by magic, appears.)
Now that we’re inside the right directory, we’re read to go! You can type “ls” to view the files in your directory. Included should be the files that you will call with the following “macqiime”commands. The program will always look for input files starting from the directory that you are in and output files will be written to this directory, so if the program can’t find files that you KNOW are there, you should make sure that you are working from the correct directory. Files you will need to get started are a mapping file “Fasting_Map.txt”, a fasta format file of sequences “Fasting_Example.fna”, and a sequence quality file “Fasting_Example.qual”.Qiime Tutorial
Now run through the Qiime 16S data analysis pipeline on your own. Refer to the online tutorial for more background and further options: http://qiime.org/tutorials/tutorial.html. You can copy and paste the following commands in your terminal prompt to run through this tutorial.
1) “Check mapping file”
macqiime check_id_map.py -m Fasting_Map.txt -o mapping_output -v
2) “Demultiplexing”
macqiime split_libraries.py -m Fasting_Map.txt -f Fasting_Example.fna -q Fasting_Example.qual -o split_library_output
3) “Pick OTUs through OTU table”
macqiime pick_otus_through_otu_table.py -i split_library_output/seqs.fna -o otus
4) “OTU Heatmap”
macqiime make_otu_heatmap_html.py -i otus/otu_table.txt -o otus/OTU_Heatmap/
5) “OTU Network”
macqiime make_otu_network.py -m Fasting_Map.txt -i otus/otu_table.txt -o otus/OTU_Network
6) “Make Taxa Summary Charts“
macqiime summarize_taxa_through_plots.py -i otus/otu_table.txt -o wf_taxa_summary -m Fasting_Map.txt
You will need to create a .txt file for the following rarefaction script. To do this, open up TextWrangler and type:
alpha_diversity:metrics shannon,PD_whole_tree,chao1,observed_species
In the same directory save this file as: alpha_params.txt
[with Unix (LF) Line breaks and Encoding with Unicode (UTF-8)]
7) “Alpha rarefaction”
macqiime alpha_rarefaction.py -i otus/otu_table.txt -m Fasting_Map.txt -o wf_arare/ -p alpha_params.txt -t otus/rep_set.tre -a
8) “Beta diversity and plots”
macqiime beta_diversity_through_plots.py -i otus/otu_table.txt -m Fasting_Map.txt -o wf_bdiv_even146/ -t otus/rep_set.tre -e 146 -a
9) “Jackknifed beta diversity”
macqiime jackknifed_beta_diversity.py -i otus/otu_table.txt -t otus/rep_set.tre -m Fasting_Map.txt -o wf_jack -e 110 -a
10) “Make Bootstrapped Tree”
macqiime make_bootstrapped_tree.py -m
11) “Make Bi-Plots”
macqiime make_3d_plots.py -i wf_bdiv_even146/unweighted_unifrac_pc.txt -m Fasting_Map.txt -t wf_taxa_summary/otu_table_L3.txt –n_taxa_keep 5 -o 3d_biplot
* The above is modified from Anthony Amends MOTHUR Tutorial for Fungal Community Analysis and Qiimes overview tutorial on 16S data.