Cell culture-based shark karyotyping as a resource for chromosome-scale genome analysis

Abstract

Karyotyping is indispensable for validating genome assemblies whose sequence lengths can be scaled up to chromosome sizes using modern methods and is traditionally performed using cytogenetic techniques. Karyotype reports of chondrichthyans are scarce, mainly because of their unique osmoregulatory mechanism, which hinders cell culture. Here, we focused on carpet shark species and the culture conditions for fibroblasts and lymphocytes. Using this method, we performed high-fidelity characterization of their karyotypes, namely 2n = 102 for the whale shark (Rhincodon typus) and zebra shark (Stegostoma fasciatum), and 2n = 106 for the brownbanded bamboo shark (Chiloscyllium punctatum) and whitespotted bamboo shark (C. plagiosum). We identified heteromorphic XX/XY sex chromosomes for the two latter species and demonstrated the first-ever fluorescence in situ hybridization of shark chromosomes prepared from cultured cells. Our technical solution is applicable to diverse chondrichthyan species and will deepen the understanding of early vertebrate evolution at the molecular level.

In vitro and in vivo gene introduction in the cloudy catshark (Scyliorhinus torazame), a cartilaginous fish

“…On the other hand, our time-lapse recordings rarely showed cell division, which is usually observed in primary cultured cells from embryos (Additional file 1). Although further quantitative examination using BrdU is necessary to confirm this, the frequency of cell division might be lower in our experiments than it has been in reports using other vertebrates including sharks [31] [32]. One possible reason for this low frequency of cell division is the low incubation temperature.…”