Each species has a typical karyotype, which represents the phenotypic appearance of the somatic chromosomes including number, size, and morphology. An idiogram is a diagrammatic representation of the chromosomes showing their relative size, homologous groups, and different cytogenetic landmarks. Chromosomal analysis of cytological preparations is an essential component of many investigations, which involves the calculation of karyotypic parameters and the generation of idiograms. Although various tools are available for karyotype analysis, here we demonstrate karyotype analysis using our recently developed tool named KaryoMeasure. KaryoMeasure is a semi-automated free and user-friendly karyotype analysis software that facilitates data collection from different digital images of metaphase chromosome spreads and calculates a wide variety of chromosomal and karyotypic parameters along with the related standard errors. KaryoMeasure draws idiograms of both diploid and allopolyploid species into a vectorbased SVG or PDF image file.