FieldLabComparison

Scripts for microbiome analysis of bacterial 16S sequences from ants.

Data:

• SRA accessions:
  • PRJNA170251 (Mycocepurus): metadata in SRA correct, data/SraRunTableSRP018247.txt
  • PRJNA170250 (Trachymyrmex/Cyphomyrmex): metadata in SRA is not correct, revised metadata in data/SraRunTableSRP018246corrected.txt
• Silva database:
  • Use Full length sequences and taxonomy references for version 128, which includes taxonomy file. The silva.nr_v128.tax file and silva.nr_v128.align file need to be moved to the silva/. Another option is to use the silva.bacteria.fasta which is a silva reference file from the MiSeq SOP. The silva.bacteria.fasta file can be trimmed to the V1-V3 region using the align.seqs() and the E.coliV1V3.fas file.

Dependencies:

• SRA Toolkit v2.8.1or higher for data download
• seq_crumbs only if using dataCheck.sh
• MothuR for raw data cleaning and OTU assessment

Analysis Design

• MothuR analysis based on 454 and MiSeq tutorials to create OTU table
• Visualization and hypothesis testing in R

Workflow

• dataDownload.sh download data from NCBI SRA (takes several minutes depending on internet connectivity)
  • data/dataCheck.sh check SRA data against archived sequence files, do not need to run again
• mothur_SilvaRef.bat create custom Silva alignment for reference (takes ~10 minutes)
• mothur_prep.sh split, trim, and aggregate sequence files (takes several minutes)
• mothur_otu.bat processes combined sequences and outputs OTU table