In vivo and in vitro development of Tibetan antelope(Pantholops hodgsonii)interspecific cloned embryos

The Tibetan antelope is endemic to the Tibetan Plateau,China,and is now considered an endangered species.As a possible rescue strategy,the development of embryos constructed by interspecies somatic cell nuclear transfer(iSCNT)was examined.Tibetan antelope fibroblast cells were transferred into enucleated bovine,ovine and caprine oocytes.These cloned embryos were then cultured in vitro or in the oviducts of intermediate animals.Less than 0.5%of the reconstructed antelope-bovine embryos cultured in vitro developed to the blastocyst stage.However,when the cloned antelope-bovine embryos were transferred to caprine oviducts,about 1.6%of the embryos developed to the blastocyst stage.In contrast,only 0.7%of the antelope-ovine embryos developed to the morula stage and none developed to blastocysts in ovine oviducts.The treatment of donor cells and bovine oocytes with trichostatin A did not improve the embryo development even when cultured in the oviducts of ovine and caprine.When the antelope-bovine embryos,constructed from oocytes treated with roscovitine or trichostatin A,were cultured in rabbit oviducts 2.3%and 14.3%developed to blastocysts,respectively.It is concluded that although some success was achieved with the protocols used,interspecies cloning of Tibetan antelope presents difficulties still to be overcome.The mechanisms resulting in the low embryo development need investigation and progress might require a deeper understanding of cellular reprogramming.

Matrix attachment regions included in a bicistronic vector enhances and stabilizes follistatin gene expressions in both transgenic cells and transgenic mice

In the present study, follistatin(FST) gene expression vectors with either a bicistronic gene transfer cassette alone, or a bicistron gene cassette carrying a matrix attachment region(MAR) were constructed and transfected to bovine fetal fibroblasts. Evaluations of both the integration and expression of exogenous FST indicated that the p MAR-CAG-FST-IRES-Ac GFP1-poly A-MAR(pMAR-FST) vector had higher capacity to form monoclonal transgenic cells than the vector without MAR,though transient transfection and integration efficiency were similar with either construct. Remarkably, protein expression in transgenic cells with the p MAR-FST vector was significantly higher than that from the bicistronic vector. Exogenous FST was expressed in all of the p MARFST transgenic mice at F_0, F_1 and F_2. Total muscle growth in F_0 mice was significantly greater than in wild-type mice,with larger muscles in fore and hind limbs of transgenic mice. p MAR-FST transgenic mice were also found with more evenly distributed muscle bundles and thinner spaces between sarcolemma, which suggests a correlation between transgene expression-associated muscle development and the trend of muscle growth. In conclusion, a p MAR-FST vector, which excluded the resistant genes and frame structure, enhances and stabilizes FST gene expressions in both transfected cells and transgenic mice.

The ecological adaptability of cloned sheep to free-grazing in the Tengger Desert of Inner Mongolia,China

Since the birth of the first cloned sheep,somatic cell nuclear transfer technology has been successfully used to clone a variety of mammals.Cloned livestock have no apparent health risks,and the quality and safety of the cloned animal products are similar to non-cloned animals.The social behavior and environmental adaptability of postnatal cloned animals,especially when used for grassland farm production purposes,is unknown.In the present study,the cloned Dorper sheep equipped with GPS location devices were free-grazed in a harsh natural environment similar to conditions commonly experienced by Mongolian sheep.The main findings of this research were as follows.(1)Under free-grazing conditions,the cloned sheep showed excellent climatic and ecological adaptability.In extreme temperature conditions ranging from–30 to 40°C,the cloned sheep maintained acceptable body condition and behaved as other sheep.(2)The cloned sheep quickly adapted from a herd feeding strategy to the harsh environment and quickly exhibited a grazing regimen as other free-grazing sheep.(3)The cloned sheep exhibited free-grazing patterns and social behavior as other sheep.(4)The cloned sheep in the harsh environment thrived and produced healthy lambs.Overall,the cloned Dorper sheep exhibited excellent ecological adaptation,which is an important consideration for breeding meat sheep by cloning.The Dorper sheep readily adapted to the free-grazing conditions on the Mongolian plateau grassland,which attests to their ability to withstand harsh environmental conditions.