primers4clades

primers4clades designs PCR primers for cross-species amplification of non-aligned protein-coding genes to target specific phylogenetic clades in metagenomic or uncharacterized organisms.

Key Features:

• Input handling: Accepts non-aligned protein-coding gene sequences that may include introns.
• Sequence alignment and phylogeny: Performs multiple sequence alignment and constructs a neighbor-joining phylogenetic tree from the alignments.
• Extended CODEHOP strategy: Implements an extended Consensus-Derived Oligonucleotide Hybridization Primers (CODEHOP) approach using both DNA and protein multiple sequence alignments.
• Thermodynamic evaluation: Evaluates thermodynamic properties of oligonucleotide pairs, including melting temperature and stability.
• Phylogenetic information content: Computes phylogenetic information content of predicted amplicons using Shimodaira-Hasegawa-like branch support values derived from maximum likelihood phylogenies.
• Non-redundant primer formulations: Produces a non-redundant set of primer formulations ranked according to thermodynamic properties.
• Amplicon distribution map: Generates an amplicon distribution map describing coverage across the target locus.

Scientific Applications:

• Metagenomics: Enables targeted amplification of novel sequences from complex environmental DNA samples to recover genes from diverse organisms.
• Evolutionary biology: Supports design of primers to capture phylogenetically informative loci for comparative and phylogenetic analyses.
• Molecular ecology: Facilitates amplification of markers from poorly characterized taxa to assess genetic diversity and species distributions.

Methodology:

Accepts non-aligned protein-coding genes (may include introns), performs multiple sequence alignment, constructs a neighbor-joining tree, applies an extended CODEHOP strategy using DNA and protein MSAs, evaluates oligonucleotide thermodynamic properties (melting temperature and stability), computes amplicon phylogenetic information using Shimodaira-Hasegawa-like branch support from maximum likelihood phylogenies, returns non-redundant primer formulations ranked by thermodynamics, and generates an amplicon distribution map.