TY - JOUR
T1 - Factors influencing the efficiency of generating genetically engineered pigs by nuclear transfer
T2 - Multi-factorial analysis of a large data set
AU - Kurome, Mayuko
AU - Geistlinger, Ludwig
AU - Kessler, Barbara
AU - Zakhartchenko, Valeri
AU - Klymiuk, Nikolai
AU - Wuensch, Annegret
AU - Richter, Anne
AU - Baehr, Andrea
AU - Kraehe, Katrin
AU - Burkhardt, Katinka
AU - Flisikowski, Krzysztof
AU - Flisikowska, Tatiana
AU - Merkl, Claudia
AU - Landmann, Martina
AU - Durkovic, Marina
AU - Tschukes, Alexander
AU - Kraner, Simone
AU - Schindelhauer, Dirk
AU - Petri, Tobias
AU - Kind, Alexander
AU - Nagashima, Hiroshi
AU - Schnieke, Angelika
AU - Zimmer, Ralf
AU - Wolf, Eckhard
N1 - Funding Information:
We are grateful to Tuna Guengoer, Eva-Maria Jemiller, Christian Erdle and Sigfried Elsner for their excellent technical support. This work was financially supported by the German Research Council (FOR 535 ‘Xenotransplantation’, FOR 793 ‘Mechanisms of Fracture Healing in Osteoporosis’, Transregio-CRC 127 ‘Biology of xenogeneic cell, tissue and organ transplantation – from bench to bedside’), by the Federal Ministry for Education and Research (Leading-Edge Cluster ‘m4 – Personalised Medicine and Targeted Therapies’), the Bavarian Research Council (FORZebRA, Az. 802–08), the Mildred Scheel Stiftung für Krebsforschung, the Mukoviszidose Institut gemeinnützige Gesellschaft für Forschung und Therapieentwicklung mbH and by the DFG International Research Training Group (1563/1 RECESS). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
PY - 2013/5/20
Y1 - 2013/5/20
N2 - Background: Somatic cell nuclear transfer (SCNT) using genetically engineered donor cells is currently the most widely used strategy to generate tailored pig models for biomedical research. Although this approach facilitates a similar spectrum of genetic modifications as in rodent models, the outcome in terms of live cloned piglets is quite variable. In this study, we aimed at a comprehensive analysis of environmental and experimental factors that are substantially influencing the efficiency of generating genetically engineered pigs. Based on a considerably large data set from 274 SCNT experiments (in total 18,649 reconstructed embryos transferred into 193 recipients), performed over a period of three years, we assessed the relative contribution of season, type of genetic modification, donor cell source, number of cloning rounds, and pre-selection of cloned embryos for early development to the cloning efficiency.Results: 109 (56%) recipients became pregnant and 85 (78%) of them gave birth to offspring. Out of 318 cloned piglets, 243 (76%) were alive, but only 97 (40%) were clinically healthy and showed normal development. The proportion of stillborn piglets was 24% (75/318), and another 31% (100/318) of the cloned piglets died soon after birth. The overall cloning efficiency, defined as the number of offspring born per SCNT embryos transferred, including only recipients that delivered, was 3.95%. SCNT experiments performed during winter using fetal fibroblasts or kidney cells after additive gene transfer resulted in the highest number of live and healthy offspring, while two or more rounds of cloning and nuclear transfer experiments performed during summer decreased the number of healthy offspring.Conclusion: Although the effects of individual factors may be different between various laboratories, our results and analysis strategy will help to identify and optimize the factors, which are most critical to cloning success in programs aiming at the generation of genetically engineered pig models.
AB - Background: Somatic cell nuclear transfer (SCNT) using genetically engineered donor cells is currently the most widely used strategy to generate tailored pig models for biomedical research. Although this approach facilitates a similar spectrum of genetic modifications as in rodent models, the outcome in terms of live cloned piglets is quite variable. In this study, we aimed at a comprehensive analysis of environmental and experimental factors that are substantially influencing the efficiency of generating genetically engineered pigs. Based on a considerably large data set from 274 SCNT experiments (in total 18,649 reconstructed embryos transferred into 193 recipients), performed over a period of three years, we assessed the relative contribution of season, type of genetic modification, donor cell source, number of cloning rounds, and pre-selection of cloned embryos for early development to the cloning efficiency.Results: 109 (56%) recipients became pregnant and 85 (78%) of them gave birth to offspring. Out of 318 cloned piglets, 243 (76%) were alive, but only 97 (40%) were clinically healthy and showed normal development. The proportion of stillborn piglets was 24% (75/318), and another 31% (100/318) of the cloned piglets died soon after birth. The overall cloning efficiency, defined as the number of offspring born per SCNT embryos transferred, including only recipients that delivered, was 3.95%. SCNT experiments performed during winter using fetal fibroblasts or kidney cells after additive gene transfer resulted in the highest number of live and healthy offspring, while two or more rounds of cloning and nuclear transfer experiments performed during summer decreased the number of healthy offspring.Conclusion: Although the effects of individual factors may be different between various laboratories, our results and analysis strategy will help to identify and optimize the factors, which are most critical to cloning success in programs aiming at the generation of genetically engineered pig models.
KW - Knockout pig
KW - Multi-factorial analysis
KW - Somatic cell nuclear transfer
KW - Transgenic pig
UR - http://www.scopus.com/inward/record.url?scp=84877857083&partnerID=8YFLogxK
U2 - 10.1186/1472-6750-13-43
DO - 10.1186/1472-6750-13-43
M3 - Article
C2 - 23688045
AN - SCOPUS:84877857083
SN - 1472-6750
VL - 13
JO - BMC Biotechnology
JF - BMC Biotechnology
M1 - 43
ER -