# A new molecular marker including parts of conservative histone H3 and H4 genes and the spacer between them for phylogenetic studies in dragonflies (Insecta, Odonata), extendable to other organisms

## TL;DR

This paper introduces a new molecular marker combining histone H3 and H4 genes and their spacer for phylogenetic studies in dragonflies and other organisms.

## Contribution

A novel histone H3–H4 marker is proposed for phylogenetic analysis at short evolutionary distances in insects and other organisms.

## Key findings

- The histone H3–H4 marker provides good resolution at family, genus, and species levels in Odonata.
- The non-coding spacer between H3 and H4 genes offers sufficient variation for short evolutionary distance analysis.
- The marker shows a nested relationship between Sympetrum croceolum and S. uniforme species.

## Abstract

In spite of recent substantial progress in genomic approaches, there is still a need for molecular markers convenient for Sanger sequencing and providing good phylogenetic reconstructions at short evolutionary distances. A new molecular marker, the histone H3–H4 region, containing partial coding sequences of the genes for histones H3 and H4 and the non-coding spacer between them, is proposed. This marker is potentially useful for molecular phylogenetic studies at the generic, species, and even intra-species level in insects and some other organisms, even from other phyla. The highly conserved histone-coding sequences ensure the universality of primers and the ease of primary alignment, while the highly variable non-coding spacer provides enough variation for analyses at short evolutionary distances. In insects, the histone genes reside in the histone repeat which is tandemly repeated in dozens to hundred copies forming the so-called histone cluster. This ensures a high concentration of the template for the marker in genomic DNA preparations. However, the order and orientation of the histone genes in the histone repeat is variable among orders, which puts some limitations on the use of the proposed marker. The marker efficacy is hereby shown for Odonata (dragonflies and damselflies), where it provided good resolution at the family, genus and species levels. The new marker also provided an interesting pattern in the relationship of two Sympetrum species, S. croceolum and S. uniforme, showing the sequences of the latter as a branch nested among those of the former. The same combination of the proposed original primers should also work in Diptera