Goat Embryo Development from in vitro Matured Oocytes of Heterogeneous Quality Through Intracytoplasmic Sperm Injection Technique

Experiments were conducted to develop an intracytoplasmic sperm injection (ICSI) protocol to produce goat embryos from in vitro matured heterogeneous oocytes. Two different protocols ('Protocol 1' or P-1 and 'Protocol 2 super(1) or P-2) were formulated and tested to develop a suitable one. Cumulus-oocyte complexes (COCs) were obtained either by LOPU (P-1) or by ovariectomy and slicing (P-2). Recovered COCs were cultured in a microdrop (50 mu L) of HEPES-buffered in vitro maturation (IVM) medium with 10% oestrus goat serum (OGS) (P-1) or IVM medium without HEPES with 15% OGS (P-2) in presence of CO sub(2)(5%) at 38.5 degree C either for 24 h (P-1) or 27 h (P-2). After IVM, COCs were denuded to select a mature oocyte (MII) and held in the incubator in presence of CO sub(2)(5%) until ICSI. Motile sperm were selected from frozen-thawed buck semen by swim-up either in mDM (P-1) or mSOF (P-2) medium. A motile sperm was immobilized and injected, head-first, into the ooplasm of a MII oocyte. Sham injections were also performed as control. Injected oocytes were cultured in mSOF medium either with 10% (P-1) or 20% (P-2) goat serum in presence of CO sub(2)(5%) at 38.5 degree C. A significantly higher (p