Recently augmentation of clinical and therapeutically important genes within the same species has become a subject of extensive research. Stearoyl-CoA desaturase (SCD) is a crucial enzyme converting saturated fatty acids to PUFAs and MUFAs and producing Conjugated Linoleic Acid (CLA) in its pathway, thus offering a complete package in granting nutritional values and immunity from fatal diseases. We aimed to identify the stable transfection method for transfecting a SCD construct in buffalo fetal fibroblast cells and determining its counter-effect on other cellular functions. Transfection of bovine SCD was performed using Nucleofection, Lipofectamine 3000 and Fugene HD. The level of transfection was analysed using flow-cytometry and efficiency was determined by MTT, relative proliferation, growth curve analysis and karyotyping. To understand the underlying genetic effect of transfection we studied the genes related to pluripotency (OCT4, SOX2, NANOG), epigenetics (DNMT1, DNMT3B, HDAC1), apoptosis (P53, CASPASE3, BAX, BID) and growth (G6PD, IGF1R). Considering the parameters tested a final transfection efficiency of 30% by nucleofection in buffalo fetal fibroblasts was observed, which was superior to all non-viral transfection methods tested to date. Also, it was observed that although all transfected cells showed higher level apoptosis, the level in nucleofected cells was the lowest. Overall when comparing the results of FACS, MTT assay and Gene expression profiling, it is suggested that nucleofection has a higher transfection efficiency and serves as a worthy candidate in delivering the expression vector with a comparatively lesser side effect on cellular functions and these cells may be further used for the production of transgenic buffalo.
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