Gene delivery to Nile tilapia cells for transgenesis and the role of PI3K-c2α in angiogenesis

Abstract

Microinjection is commonly performed to achieve fish transgenesis; however, due to difficulties associated with this technique, new strategies are being developed. Here we evaluate the potential of lentiviral particles to genetically modify Nile tilapia cells to achieve transgenesis using three different approaches: spermatogonial stem cell (SSC) genetic modification and transplantation (SC), in vivo transduction of gametes (GT), and fertilised egg transduction (ET). The SC protocol using larvae generates animals with sustained production of modified sperm (80% of animals with 77% maximum sperm fluorescence [MSF]), but is a time-consuming protocol (sexual maturity in Nile tilapia is achieved at 6 months of age). GT is a faster technique, but the modified gamete production is temporary (70% of animals with 52% MSF). ET is an easier way to obtain mosaic transgenic animals compared to microinjection of eggs, but non-site-directed integration in the fish genome can be a problem. In this study, PI3Kc2α gene disruption impaired development during the embryo stage and caused premature death. The manipulator should choose a technique based on the time available for transgenic obtainment and if this generation is required to be continuous or not.

Hybrid Sterility in Fish Caused by Mitotic Arrest of Primordial Germ Cells

“…At present, several different approaches to germ cell transplantation techniques, either within the same species (allogenic) or between two different species (xenogenic), have been established in fish (Yoshizaki et al 2011;Lacerda et al 2013) and are widely used to study germ cell biology, such as the function of germline stem cells (Yoshizaki et al 2012;Hayashi et al 2014). These techniques also have applications in the conservation of endangered species (Lee et al 2015;Pšenička et al 2015;Lee and Yoshizaki 2016;Ye et al 2017), the propagation of valuable fishes with economically useful traits (Morita et al 2012(Morita et al , 2015Silva et al 2016), the cryopreservation of scientific bioresources (Lee et al 2013(Lee et al , 2016aSeki et al 2017), and the efficient production of genetically engineered fish (Tonelli et al 2017). For the purpose of germ cell transplantation experiments, sterilized recipients are useful for obtaining successful colonization of donor germ cells in recipient genital ridges and gonads, and the efficient production of donor-derived gametes without contamination of recipient-derived gametes.…”

Transgenic Rodent Models in Toxicological and Environmental Research: Future Perspectives

“…Frogs 45,46 Fish Zebra fish 47,48,49 Altlantic salmon 50,51 Channel catfish 52,53 Goldfish 54 Nile thilapia 55,56 Invertebrates Silkworms 57,58 Nematode 59 therapeutic mAbs, including two diagnostic mAb. 64 Recently (December 24, 2021) US-FDA approved mAb (EVUSHELD™ [tixagevimab co-packaged with cilgavimab]) for preexposure prophylaxis (prevention) of COVID-19.…”

Bigger, stronger, better: Fish transgenesis applications and methods

“…When retroviruses and lentiviruses are internalized into the target cell, their genetic material (RNA) is transformed to DNA, thereby their genetic information is inserted into the host genome (Tonelli et al, 2017a). After transduction efficiency of pseudotyped viruses was displayed on zebrafish in 1994, this method was applied on various fish species including nile tilapia and live-bearing fish (Poeciliposis lucida) to integrate transgene into the fish genome (Lin et al, 1994;Sarmasik et al, 2001;Tonelli et al, 2017b).…”