39 FACTORS INFLUENCING THE EFFICIENCY OF GENERATING GENETICALLY ENGINEERED PIGS BY NUCLEAR TRANSFER: MULTI-FACTORIAL ANALYSIS OF A LARGE DATA SET
M. Kurome A , L. Geistlinger B , B. Kessler A , V. Zakhartchenko A , N. Klymiuk A , A. Wuensch A , K. Flisikowski C , T. Flisikowska C , C. Merkl C , H. Nagashima D , A. Schnieke C , R. Zimmer B and E. Wolf AA Chair for Molecular Animal Breeding and Biotechnology, and Laboratory for Functional Genome Analysis (LAFUGA), Gene Center, LMU Munich, Munich, Germany;
B Chair for Practical Informatics and Bioinformatics, Institute for Informatics, LMU Munich, Munich, Germany;
C Chair for Livestock Biotechnology, Center of Life and Food Sciences Weihenstephan, TU Munich, Freising, Germany;
D Meiji University, Kawasaki, Japan
Reproduction, Fertility and Development 25(1) 167-167 https://doi.org/10.1071/RDv25n1Ab39
Published: 4 December 2012
Abstract
Somatic cell nuclear transfer (SCNT) using genetically engineered donor cells is currently the most widely used strategy to generate genetically tailored pig models for biomedical research. Although this approach facilitates a similar spectrum of genetic modifications as in rodent models, including inducible transgene expression and gene targeting, the outcome in terms of live cloned piglets is quite variable. We used a large data set from 274 SCNT experiments (in total, 18 649 reconstructed embryos transferred into 193 recipients), performed over a period of 3 years, and evaluated it by multivariate analysis for combined assessment of multiple factors and their relative contribution to the efficiency of generation of genetically modified cloned pigs. Specifically, we addressed the effects of season, type of genetic modification (additive gene transfer v. gene targeting), donor cell source (mesenchymal stem cells, postnatal fibroblasts, fetal fibroblasts, and kidney cells), serial NT, and pre-selection of SCNT embryos for early development. Target parameters were pregnancy and delivery rates, as well as the numbers of born, live, and healthy offspring. Cloning efficiency was calculated as the number of cloned piglets relative to the number of transferred SCNT embryos. We used robust linear models adjusted to the underlying empirical distribution of the cloning outcome, as a straightforward approach to determine the statistically significant part of the network of factors affecting pig cloning. The SCNT experiments performed during the winter using fetal fibroblasts or kidney cells after additive gene transfer resulted in the highest number of live and healthy offspring, whereas sequential NT and NT experiments performed during the summer decreased it. Although the effects of individual factors may be different between various laboratories, the statistical approach described in this study may help to identify and optimize the most critical specific factors to cloning success in programs aimed at the generation of genetically engineered pig models.
Supported by the DFG (FOR535, FOR793), the Bayerische Forschungsstiftung, and Mukoviszidose e.V.
