Pipeline of the MicroSyn. In the gene list file, one line contains chromosome or fragment name, gene accession number, the '+' or '-' sign indicating the orientation of the gene on strand, and the absolute or relative position. Gene sequence file contains corresponding gene's CDS in FASTA format. These two files and genes in a family are merged into a file, called MSY file here. After loading the MSY file, MicroSyn then initially creates a homologous relationship among all genes on fragments and a table consisting of all pairs of homologous genes is obtained. Next, MicroSyn begins detecting microsynteny between fragments that contain the members in a gene family. Finally, the software also estimates the level of Ks of paralogs between two fragments, using the flanking gene sequences.