Abstract Two correlated genetic features are characteristic of the order Hymenoptera, i.e., arrhenotoky and haplodiploidy, but multiple transitions to diploid thelytoky also occurred within this group. Karyotypes of approximately two thousand members of the order are recently known. History of the chromosomal study of the Hymenoptera can be provisionally subdivided into four stages, with approximate borders of the 1930s, 1970s and 2000s between them. Although the development of this study can mainly be explained by the technical progress in preparing and analyzing chromosomal preparations, the results obtained with the help of earlier developed methods, also can successfully be used nowadays. In addition to morphometric analysis, a number of differential staining techniques are used to identify particular chromosomes and their segments; these techniques can conditionally be subdivided into two groups, the so-called “traditional” and “modern” ones. First of all, C- and AgNOR-bandings constitute the former methods; these techniques visualize heterochromatic segments and nucleolus organizing regions respectively. Moreover, modern methods are also widely used at present for studying parasitoid karyotypes. These techniques include use of fluorescent dyes (fluorochromes), especially those specifically staining AT- and GC-rich chromosome segments. Fluorescence in situ hybridization (FISH) is a very important method of physical mapping of DNA sequences on chromosomes. Immunocytochemical techniques can be of use to study chemical content and structure of chromosomes; these methods involve use of specific fluorochrome-conjugated antibodies. Nowadays, taxonomic significance of karyotypic study of the order Hymenoptera substantially increases, especially within the framework of the so-called integrative taxonomy, aimed for recognition, delimitation and description of closely related species. Furthermore, a combined use of classical and molecular methods has very good perspectives. Knowledge of hymenopteran phylogeny is necessary for identifying pathways of karyotype evolution of the order, but at least in some cases chromosome characters can be considered as synapomorphies defining different lineages. Karyotypic research also has very important implications for genetic studies of Hymenoptera. The chromosome number equals the number of linkage groups within the genome, but it also can be used as a proxy to the level of genetic recombination, especially in the context of big data approach. In addition, significance of physical mapping of DNA sequences increases in the light of the modern efforts in genome sequencing. FISH is most often used for mapping repetitive sequences, including ribosomal DNA, microsatellites and telomeric segments. Nevertheless, this technique could be useful for mapping unique sequences as well. In the order Hymenoptera, FISH is also successfully used together with chromosome microdissection for identifying particular chromosomes and/or chromosome segments, as well as various chromosomal rearrangements. In addition, chromosomal analysis can reveal the so-called supergenes, i.e., inverted chromosome segments, which accumulate genetic differences. Finally, immunocytochemical techniques can map distribution of various chemical compounds along the chromosomes, including identification of the degree of methylation of the chromosomal DNA.
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