Effect of custom-designed transfection buffer on delivery of genome modification components into primary cells of buffalo, cattle, goats, and sheep
Abstract
The transfer of genome-modification components into farm animal cells is indispensable for the production of genome-modified and transgenic farm animals. Electroporation is a physical transfection method when appropriately used; this technique is safe, simple to use, affordable, and efficient in transfecting cells from several lineages. Electroporation efficiency depends on various physical parameters, of which cell type is considered a major factor for transfection efficiency. Primary cells are generally less susceptible to transfection than other cell types due to their finite lifespan and limited expansion capacity. Previously, we custom-designed a transfection buffer to deliver exogenous genetic components into mammalian cells. In the present study, we examined the effect of the developed buffer on transfection rates and cell viability of primary somatic cells from buffalo, cattle, goats, and sheep. To achieve the aims of this study, t primary somatic cells from skin biopsies were established and were transfected with a Venus-expression vector (pAcGFPs-Venus). We noticed that transfection rates of pAcGFPs-Venus were 22.51%, 17.56%, 22.81%, and 16.16% for buffalo, cattle, goats, and sheep cells, respectively. We also noticed that cell viability and proliferation rates were better in the case of goats, sheep, and cattle cells; also, these cells have less vacuolation than buffalo cells. In addition, we also generated MSTN (myostatin) KO (Knockout) cell clones from these cell populations, in which the efficiency of single-cell clone generation was high for goats and sheep cells. In conclusion, our lab-made transfection buffer can be efficiently used to generate genome-edited or transgenic farm animals for agriculture, biomedical, and veterinary applications.
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