DeGenPrime

DeGenPrime designs PCR primers by utilizing thermodynamic properties, filtration metrics, and conserved region finding to expand the target range for a single primer set.

Key Features:

• MSA format support: DeGenPrime accepts multiple sequence alignment formats as input for primer design.
• Degenerate base handling: The tool incorporates degenerate bases to expand primer target range across diverse sequences.
• Degeneracy filter: A filtration metric controls the level of degeneracy permitted in designed primers.
• Repeated k-mers filter: The tool filters primers based on repeated k-mer content.
• Relative GC content filter: GC content is evaluated and filtered as a primer quality metric.
• Temperature range filter: Primers are filtered according to specified temperature range parameters.
• Minimal penalty scoring: The tool applies penalty scoring based on secondary structure self-dimerization metrics, GC clamping, tri-loop hairpins, tetra-loop hairpins, and internal repetition.

Scientific Applications:

• Alternative nitrogenase gene targeting: DeGenPrime successfully designed primers for the alternative iron nitrogenase (anfD) gene target where PrimerDesign-M, DegePrime, and ConsensusPrimer achieved 0%, 11%, and 17% acceptable primer yield, respectively.
• T4-like phage major capsid protein analysis: The tool identified quality primers within conserved regions of the T4-like phage major capsid protein (g23).
• Nitrogenase family primer design: DeGenPrime designed primers for conserved regions of molybdenum-based nitrogenase (nif) and its alternatives vanadium (vnf) and iron (anf) nitrogenase.
• Universal primer design: The tool provides scalable primer design applicable across the entire tree of life.

Methodology:

DeGenPrime utilizes nearest-neighbor thermodynamic properties, conserved region finding, and filtration via penalty of oligonucleotides. Penalty scoring incorporates secondary structure self-dimerization metrics, GC clamping, tri-loop hairpins, tetra-loop hairpins, and internal repetition. The tool is written in C++.