An efficient protocol for chromosome isolation from sterlet (A. ruthenus) embryos and larvae

Abstract

In the present manuscript an efficient and feasible protocol for chromosome preparation from sterlet (A. ruthenus) embryos and larvae was developed that can be used as routine molecular diagnostic tool of the species genome manipulation procedures verification. For this end, a multi-dimensional experimental system was conceived that enabled for the establishment of chromosome preparation protocol, which characterized the mean efficiency of chromosome extraction ranged from 70% to 100% and the average number of recorded metaphases per slide ranged between 9 and 15. The developed under the current study protocol can be used as a fast and reliable tool, alternative for flow cytometry techniques that enable for the molecular verification of genome manipulations effectiveness. As genome engineering is presently the fundamental biotechnology tool that enables for effective and sustainable aquaculture of fish, including sturgeons that are known as one of the most valuable economically and ecologically fish group, the development of easy and fast methods for the verification of genome manipulation effects is especially important. Thus, the established chromosome preparation protocol contributes to genome and reproduction studies of sturgeons, including taxonomy, inter-species hybridization, genome aberrations, sex determination system and selection. Moreover, the application of cytogenetic techniques also contributes to the development of new or improvement existing techniques of genome engineering utilized in sturgeon aquaculture.