PriSeT

Tool for assisting the search of primer sequences for metabarcoding experiments. Given a taxonomic identifier, a reference database, and a genetic target region, PriSeT identifies conserved sections suitable for PCR primers such that taxonomic coverage and separation are optimal.

Platform	Details	Binaries
	Mac OS 64 bit	Link to Build

Usage

Usecase: Precompiled Binary

Download one of the above linked binaries (Ubuntu build will be provided soon) and call it from command line with a directory containing a FASTA file of the reference sequences. The binary is based on apps/solver_fast.cpp. In case you want to edit the app and change the primer properties via setter functions provied by types/PrimerConfig.hpp, follow the compilation instructions in the next section. The FASTA file should not be larger than 500 MB. Index building and primer search are decoupled to allow for tweaking the constraints that affect the primer pair result set.

Build FM-Index

First an FM-index is built on the FASTA file and stored under the user-given directory. Note, that the FM-index consumes about four times more space than the input file. The index for a specific library has to be built only once. Create work_dir in advance.

./priset -i -l <path_to_fasta> -w <work_dir|idx_dir>

Discover Primer Pairs

Primer search is triggered via the -s flag. The results are output in csv format; the first 20 pairs are displayed in terminal.

./priset -s -l <path_to_fasta> -w <work_dir>

Usecase: Userwritten Apps

In case you want to modify the way how primer pairs are searched and processed, you can write and compile your own applications.

Requirements

• SDSL-Lite, get it from here: xxsds/sdsl-lite
• Cmake 3.7 or higher
• GNU C++ Compiler xx or higher

Setup

Clone PriSeT and its submodules

git clone --recurse-submodules https://github.com/mariehoffmann/PriSeT.git

Compilation

User-defined apps should go into the apps folder, and the therein contained CMakeLists.txt file modified. In the following we assume your application is called solver_fast.cpp. Add the following entry to the apps/CMakeLists.txt:

priset_app_macro(solver_fast.cpp)

The macro will create a target and link it against the required libraries.

1. Create a build directory and compile with cmake

cmake ../PriSeT/apps -DCMAKE_BUILD_TYPE=Debug -B .

2. Build

make -j

3. Run to build FM-index once

./solver_fast -i -l <lib_dir> -w <work_dir>

4. Discover primers

./solver_fast -s -l <lib_dir> -w <work_dir>

Unit Tests

Add new unit tests under test/unit and run cmake

cmake ../PriSeT/test/unit/ -DGENMAP_NATIVE_BUILD=ON -DCMAKE_BUILD_TYPE=Debug -B .

References

[1] Pockrandt, C., Alzamel, M., Iliopoulos, C. S., Reinert, K.. GenMap: Fast and Exact Computation of Genome Mappability. bioRxiv, presented on RECOMB-Seq, 2019.

[2] Federhen, S. (2012). The NCBI Taxonomy database. Nucleic Acids Research, 40(D1), D136–D143. http://doi.org/10.1093/nar/gkr1178

[3] PCR primer design constraints: www.premierbiosoft.com/tech_notes/PCR_Primer_Design.html