The script has been updated to ensure smooth operation of the translation step.
Additional notes have been added (see Reporting bugs section below) to solve possible issues during PANDASeq installation in Mac computers with Apple silicon.
This is the README document for the EasyDIVERS pipeline for pre-processing HTS reads from in vitro selection experiments. The pipeline can be used to process nucleotides or amino acids sequencing data.
Please consult the EasyDIVER manual.
easydiver -i [-o -p -q -T -a -r -e -h]
| Flag | Description | Comments |
|---|---|---|
| -i | Input directory path and name | Required |
| -o | Output directory path and name | Optional Default value: /pipeline.output |
| -p | Extraction forward DNA primer | Optional |
| -q | Extraction reverse DNA primer | Optional |
| -T | Number of threads used for computation | Optional Default value: 1 |
| -a | Translation into amino acids is performed | Optional Default value: FALSE |
| -r | Files for individual lanes are retained | Optional Default value: FALSE |
| -e | Additional internal PANDAseq flags | Optional Must be entered in quotation marks (e.g. -e “-L 50”) Default value: “-l 1 -d rbfkms“ |
| -h | Help message | Optional |
The pipeline script was written to run on Unix-based systems, like Linux, Ubuntu, and MacOS. Windows 10 also has a Linux subsystem.
To use the pipeline, first install the two dependencies: Python and PANDASeq. We recommend using the Anaconda distribution of python, and adding the Bioconda channel to Anaconda's package manager, conda. See the Anaconda documentation for installation. After installing Anaconda with Bioconda, PANDASeq is easily installed using conda with:
conda install pandaseq
In order for the pipeline to be called from any directory and for the pipeline to call the translator reliably, both scripts must be placed in a directory that is in the user's PATH environment variable upon download. For example, for Unix/Linux users, scripts could be placed in /usr/local/bin/ upon download. These files can be placed in that directory with the command:
cp /path/to/pipeline.sh /path/to/translator.py /usr/local/bin/
EasyDIVER and the translation tool must be made executable. This can be done by entering the following commands from the local directory where they are stored:
chmod +x easydiver.sh
chmod +x translator.py
The pipeline will not be found unless it is stored in the working directory or in a directory that is in the user's PATH environment (e.g. bin/). Also, the pipeline will not be able to find the translator if it is not stored in a directory that is in the user's PATH environment (e.g. bin/).
All input files must be:
- Located in the same directory (even reads from separate lanes).
- In FASTQ format
- Named using the standard Illumina naming scheme: sample-name_S#_L00#_R#_001.fastq
- In either .fastq or .fastq.gz extensions.
A test dataset is provided. The test data corresponds to two samples obtained from a real experiment of in vitro evolution of mRNA displayed peptides.
Please report any bugs to Celia Blanco (celiablanco@ucla.edu).
When reporting bugs, please include the full output printed in the terminal when running the pipeline.
If a problem is encountered with newer MacOS versions after installing PANDASeq, you may try the following:
- Install Homebrew (see here: https://brew.sh/)
- brew install bzip2 pkgconfig libtools
- Run the ./autogen.sh build step (see PANDASeq manual)
If an error referencing snprintf occurs, identify the file from the error message, open that file and adjust 'snprintf' to be 'printf' instead. During our test runs, this issue was found in line 528 in the pandaseq package args.c file. Run the ./autogen.sh build step again. At this point, you might get many ‘warnings’ but you shouldn't get any errors.
Celia Blanco*, Samuel Verbanic*, Burckhard Seelig and Irene A. Chen. EasyDIVER: a pipeline for assembling and counting high throughput sequencing data from in vitro evolution of nucleic acids or peptides. J Mol Evol 88, 477–481 (2020).