InMethyl

InMethyl is a Python-based application for the design of target-specific primer combinations for PCR amplification and bisulfite sequencing of complete CpG-islands. InMethyl uses bowtie high-throughoutput aligner to identify potential mis-priming sites in the bisulfite treated or intact genome and then – undesirable potential PCR products. InMethyl provides a balance between various characteristics that allows to pick up primers in hard-to-study genomic regions. This balance is based on the calculation of scoring factor including primer pair specificity, nucleotide composition (sequence complexity), thermodynamic features (melting temperature, dimers dG etc.), presence of CpG-sites and other parameters. Users are intended to customize the desired or limit ranges of these values, as well as penalties for out-of-bounds values. InMethyl software allows to optimize combination of PCR primer pairs to perform the amplification of large genomic regions, e.g. CpG-islands.