Robust DNase activity of the ooplasm can act as a gametic transfection barrier in rainbow trout

Barkhojasteh, Mania; Abdolhay, Hossein Ali; Gorjipoor, Einollah; Amini, Hamidreza; Salahi, Mohammad Meysam;
Department of Biotechnology, Institute of farm animal genetics (ING), Friedrich-Loeffler-Institut, Mariensee, Neustadt am Rübenberge, Germany
Eghbalsaied, Shahin

In this study, we evaluated DNase activity of rainbow trout oocyte using an in vitro and in vivo study. First, synthetic single strand and natural double strand DNA from Eukaryotic and prokaryotic sources as well as naked DNA were in vitro incubated with the oocyte cytoplasm. Results showed that the DNase activity of rainbow trout oocyte is strong enough to degrade any type of DNA at the onset of the incubation. Then, we evaluated if similar to the mammalian species, dead spermatozoa from rainbow trout can protect exogenous DNA from oocyte DNases. A series of dead spermatozoa was incubated with pDB2, carrying EGFP transgene, for 30 min followed by the ooplasm treatment for an additional 30 min. Not only did oocyte DNases completely degrade the exogenous DNA, but also it degraded the compact genome of spermatozoa. In conclusion, in vitro results clearly showed that strong DNase activity of ooplasm could degrade any types of foreign DNAs including oligonucleotides and intensively compact sperm genome. The strong DNase activity of rainbow trout ooplasm could be a stumbling block for genetic modification using plasmids in salmonids.



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