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First study of sperm mediated gene transfer in Egyptian river buffalo

Mohamed S Hassanane¹, Aida I El Makawy¹, Sahar M Helalia¹

¹Cell Biology Department, Genetic Engineering and Biotechnology Division, National Research Centre, Egypt.

Journal, Genetic Engineering & Biotechnology

|January 17, 2019

Abstract:

The present study was carried out to find the best treatments for enhancing the ration of insertion of a desired gene construct (pEGFP-N1) onto the sperm of buffalo as the first step for the production of transgenic buffalo using sperm mediated gene transfer (SMGT). The tested conditions were plasmid DNA concentration, sperm concentration, transfecting agent concentration: Dimethyle sulphoxide (DMSO) and time of transfection. The study proved that the best conditions for producing transgenic embryos were incubation sperm solution its concentration is 10⁷/ml sperm with 3% DMSO: with 20 µg/ml from the linarized DNA, for 15 min at 4 °C are the best conditions to produce transgenic buffalo embryo using sperm mediated gene transfer.

Keywords:

ANOVA, analysis of variance Buffalo embryos CCC, covalently closed circular COCs, Cumulus oocyte complexes DMSO DMSO, Dimethyle sulphoxide EGFP EGFP, enhanced green fluorescent protein IVF, in vitro fertilization MII, second meiotic division OD, optical density PBS, Phosphate buffer saline SMGT SMGT, sperm mediated gene transfer TCM199, tissue culture medium Transgenic

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