In this repository you can find scripts, data and output to analyse microbial communities (fungi and bacteria) from soil samples using eDNA meta-barcoding.
/bin contains documented bash and R scripts to quality-filter Illumina MiSeq 2x300 reads, generate OTUs, filter the OTU table, assign taxonomy and trophic mode (fungi), and analyze alpha and beta diversity patterns.
Program used:
/data contains the data and metadata associated to the project, apart from large fastq files (raw data).
/output contains the final tables and figures resulting from the scripts for each locus:
- Sequence quality check (FastQC reports)
- OTU table with raw count per sample
- Taxonomy and reference sequences of OTUs
- Tables with most frequent/abundant OTUs and fungal genera
- Diversity plots
technical_report.md discusses each step of the pipeline used to generate the OTU tables and assign functional guilds to fungal OTUs, as well as providing suggestions for future studies.
diversity_report.md provides some baseline diversity analyses and briefly discuss the results.
Soil samples were collected from 106 sites across the state of Victoria, at two soil depth (0–10 cm and 10–20 cm), for a total of 212 samples. Vegetation, edaphic variables and additional meta-data associated to the project can be found here.
Amplicons were generated, sequenced and demultiplexed by BioPlatform Australia for the following regions:
- ITS (fungi) with primers ITS1F/ITS4
- 16S (bacteria) with primers 27F/519R
The following control samples were included:
- Commercial mock communities: ATCC1010MOCK for ITS and ATCC1002MOCK for 16S
- Soil = Positive control as a mixture of various soil samples
- No = Negative control
Detailed protocols for sampling and bench work can be found here.