Factors Determining the Efficiency of Porcine Somatic Cell Nuclear Transfer: Data Analysis with Over 200,000 Reconstructed Embryos

Abstract

Data analysis in somatic cell nuclear transfer (SCNT) research is usually limited to several hundreds or thousands of reconstructed embryos. Here, we report mass results obtained with an established and consistent porcine SCNT system (handmade cloning [HMC]). During the experimental period, 228,230 reconstructed embryos and 82,969 blastocysts were produced. After being transferred into 656 recipients, 1070 piglets were obtained. First, the effects of different types of donor cells, including fetal fibroblasts (FFs), adult fibroblasts (AFs), adult preadipocytes (APs), and adult blood mesenchymal (BM) cells, were investigated on the further in vitro and in vivo development. Compared to adult donor cells (AFs, APs, BM cells, respectively), FF cells resulted in a lower blastocyst/reconstructed embryo rate (30.38% vs. 37.94%, 34.65%, and 34.87%, respectively), but a higher overall efficiency on the number of piglets born alive per total blastocysts transferred (1.50% vs. 0.86%, 1.03%, and 0.91%, respectively) and a lower rate of developmental abnormalities (10.87% vs. 56.57%, 24.39%, and 51.85%, respectively). Second, recloning was performed with cloned adult fibroblasts (CAFs) and cloned fetal fibroblasts (CFFs). When CAFs were used as the nuclear donor, fewer developmental abnormalities and higher overall efficiency were observed compared to AFs (56.57% vs. 28.13% and 0.86% vs.

CRISPR/Cas9-mediated integration of large transgene into pig potential safe harbor

“…2015). Our results also showed a similar mortality rate (KI 50% (2/4), non-KI 40% (2/5)) as previous reports (Liu et al . 2015; Schmidt et al .…”

“…2002). Therefore, cloned pigs have high deformity rate and mortality produced by SCNT, and the loss of cloned piglets is generally high (58%) before weaning (Liu et al . 2015; Schmidt et al .…”

Probability, odds and random chance: the difficult task of modulating the epigenetic profile of cloned embryos

“…Pieces of evidence suggest that in fact a passive process also occurs in the paternal genome, by the convertion of 5'-methylcytosine (5mC) into 5'-hydroximethylcytosine (5hmC) prior to demethylation per se (Wossidlo et al, 2011). Enzymes previously linked to chromosomal translocations in leukemia (Lorsbach et al, 2003), the ten-eleven translocation proteins (TET1, 2 and 3), have been recently implicated as responsible for the oxidation of 5mC into 5hmC (Kriaucionis and Heintz, 2009;Tahiliani et al, 2009). Then, the final demethylation process is resolved by enzymes of the DNA repair machinery in the cell (Fig.…”

Cloned pig fetuses exhibit fatty acid deficiency from impaired placental transport

“…Pig somatic cell nuclear transfer (SCNT) has valuable applications in agriculture, biomedicine, and life science (Matsunari & Nagashima, ; Yang & Wu, ). However, SCNT‐derived cloned pig fetuses frequently exhibit aberrant intrauterine development (Ao et al, ; Ao, Li, et al, ; Chae et al, ; Kim et al, ; Ruan et al, ), leading to high stillbirth occurrence (17–32.8%; Ao, Zhao, et al, ; Estrada et al, ; Huan et al, ; Kurome et al, ; Liu et al, ), congenital malformations (29.5–60.0%; Kurome et al, ; Schmidt, Winther, Secher, & Callesen, ), and neonatal mortality (48.0–74.5%; Ao et al, ; Park et al, ; Schmidt et al, ) in newborn cloned piglets. Nevertheless, available information regarding the causes of erroneous intrauterine development of cloned pig fetuses is extremely limited.…”