Requirements

You'll need to download or clone our main GitHub repository (https://github.com/mlangill/microbiome_helper) if you want to use our scripts.

In addition, the following programs should be installed with commands accessible from the user's PATH, before trying to run any of the scripts included in this repository.

Both pipelines

• FastQC (optional): http://www.bioinformatics.babraham.ac.uk/projects/fastqc/
• PEAR: http://sco.h-its.org/exelixis/web/software/pear/doc.html
• GNU Parallel (>= v20170322): https://www.gnu.org/software/parallel/

Metagenomics

• Bowtie2: http://bowtie-bio.sourceforge.net/bowtie2/index.shtml
• Human pre-indexed database: ftp://ftp.ccb.jhu.edu/pub/data/bowtie2_indexes/hg19.zip
• DIAMOND (> v.0.7.0): http://ab.inf.uni-tuebingen.de/software/diamond/
• MetaPhlAn2: https://bitbucket.org/biobakery/metaphlan2
• HUMAnN: http://huttenhower.sph.harvard.edu/humann

16S

• FASTX toolkit (v0.0.14): http://hannonlab.cshl.edu/fastx_toolkit/download.html
• BBMap: http://sourceforge.net/projects/bbmap
• VSEARCH: https://github.com/torognes/vsearch
• QIIME: http://qiime.org
• SortMeRNA: http://bioinfo.lifl.fr/RNA/sortmerna/
• SUMACLUST: http://metabarcoding.org/sumatra
• PICRUSt: http://picrust.github.io/picrust/

Visualization

• STAMP: http://kiwi.cs.dal.ca/Software/STAMP

Databases (dropbox download links)

• 16S chimera checking: RDP_trainset16_022016.fa (17 MB). This DB was originally from the Ribosome Database Project (RDP) and was parsed to include only bacteria.

• 18S chimera checking: Eukaryota_SILVA_123_SSURef_Nr99_tax_silva_U-replaced.fa (117 MB). This DB was taken from SILVA and was parsed to include only eukaryotes and all Us were converted to Ts.

• ITS2 chimera checking: UNITE_uchime_ITS2only_01.01.2016.fasta (8.6 MB) taken from UNITE database.

• Eukaryotic OTU-picking files:

  • gb203_pr2_all_10_28_99p_tax_Xs-fixed_poly-fixed.txt (9 MB)
  • gb203_pr2_all_10_28_99p_clean.fasta (98 MB)
  • 90_Silva_111_rep_set_euk_aligned.filter.fasta (95 MB)

• ITS2 OTU-picking files (taken from UNITE database):

  • UNITE_sh_refs_qiime_ver7_dynamic_20.11.2016.fasta (16.9 MB).
  • UNITE_sh_refs_qiime_ver7_dynamic_20.11.2016.goodASCII.txt (4.2 MB) - we converted all non-ASCII characters to ASCII format.

• To use "run_human_filter.pl" you will need to download the appropriate Bowtie2 index.

• To use "run_pre_humann.pl" you will need to use a database of KEGG orthologs. You may also want to download the raw FASTA.

Perl modules

File::Basename Getopt::Long List::Util Parallel::ForkManager Pod::Usage