The developmental potency of pre-implant parthenogentic goat embryos were compared under two chemical activation protocols in three different culture media groups. The in vitro matured oocytes were chemically activated by two protocols viz. P1 (CB-CHX-6DMAP) and P2 (Ca-CHX-6DMAP). The activated oocytes under both the protocols were developed in three culture media, viz. modified synthetic oviductal fluid (mSOF), research vitro cleave medium (RVCL), and RVCL-Blast. While comparing the developmental potential of activated oocytes, it was observed that the oocytes activated under P2 protocol pooled over three culture media group producing significantly higher mean cleavage rate (43.2±0.9 vs 40.6±1.5), blastocyst development (16.4±1.1 vs 12.6±0.8), and blastomere count (120.7±4.7 vs 113.2±4.1) as compared to P1 protocol. The comparison of effect of culture media pooled over protocol groups revealed that the mean cleavage rate observed under RVCL-Blast (44.8±1.3) and RVCL (45.3±0.5) were significantly higher (P≤0.01) than mSOF (35.8±1.2). However, the mean blastocyst development observed under RVCL-Blast group (18.8±3.2) was significantly higher than RVCL (14.0±0.8) and mSOF (10.8±0.4). Similarly, the mean blastomere count under RVCL-Blast group (136.0±3.7) was significantly higher (P≤0.01) than RVCL (114.7±1.0) and mSOF (100.2±0.5) groups. The semiquantitative RT PCR analysis showed the expression of pro-apoptotic caspase 3 gene in P1 and anti-apoptotic Mcl-1 gene in P2. This study concludes that the activation protocol P2 and embryo cultured under RVCL-Blast group were optimum for chemical activation and culture medium, respectively.
Skeletal muscle is the major component of lean tissue that is used for consumption, and myostatin is a negative regulator of skeletal muscle growth. Downregulation of this gene therefore offers a strategy for developing superior animals with enhanced muscle growth. Knockdown of myostatin was achieved by RNA interference technology. The anti-myostatin shRNA were designed and stably transfected in caprine fibroblast cells. The reduced expression of target gene was achieved and measured in clonal fibroblast cells by real-time PCR. Two single-cell clones induced significant decrease of myostatin gene expression by 73.96 and 72.66 %, respectively (P < 0.05). To ensure the appropriate growth of transfected cell, seven media were tested. The best suited media was used for transfected fibroblast cell proliferation. The findings suggest that shRNA provides a novel potential tool for gene knockdown and these stably transfected cells can be used as the donor cells for animal cloning.
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