Successful cloning of an adult breeding boar from the novel Chinese Guike No. 1 swine specialized strain

Abstract

Somatic cloning, also known as somatic cell nuclear transfer (SCNT), is a promising technology which has been expected to rapidly extend the population of elaborately selected breeding boars with superior production performance. Chinese Guike No. 1 pig breed is a novel swine specialized strain incorporated with the pedigree background of Duroc and Chinese Luchuan pig breeds, thus inherits an excellent production performance. The present study was conducted to establish somatic cloning procedures of adult breeding boars from the Chinese Guike No. 1 specialized strain. Ear skin fibroblasts were first isolated from a three-year-old Chinese Guike No. 1 breeding boar, and following that, used as donor cell to produce nuclear transfer embryos. Such cloned embryos showed full in vitro development and with the blastocyst formation rate of 18.4 % (37/201, three independent replicates). Finally, after transferring of 1187 nuclear transfer derived embryos to four surrogate recipients, six live piglets with normal health and development were produced. The overall cloning efficiency was 0.5 % and the clonal provenance of such SCNT derived piglets was confirmed by DNA microsatellite analysis. All of the cloned piglets were clinically healthy and had a normal weight at 1 month of age. Collectively, the first successful cloning of an adult Chinese Guike No. 1 breeding boar may lay the foundation for future improving the pig production industry.

Generation of transgenic‐cloned Huanjiang Xiang pigs systemically expressing enhanced green fluorescent protein

“…In the present study, we transferred the transgenic‐cloned embryos to surrogate sows for producing live transgenic‐cloned Huanjiang Xiang pigs. We finally obtained live transgenic‐cloned Huanjiang Xiang pigs with cloning efficiency only as 0.6% (3/504), which was similar to our previous works focused on cloning and genetic modification of other pig breeds (Liu et al, ; Nie et al, ; Zhu, Quan, et al, ). Such low efficiency (<1%) of pig cloning could be largely attributed from the poor quality of the SCNT‐derived embryos.…”

“…In vitro maturated porcine oocytes were used as nuclear transfer recipients and prepared according to described previously (Nie et al, ; Zhu, Nie, et al, ). In brief, cumulus–oocyte complexes (COCs) were aspirated from the follicles and washed twice in TL‐HEPES medium supplemented with 0.01% polyvinyl acetate (PVA).…”

“…Somatic cell nuclear transfer was performed as described previously with minor modification (Nie et al, ; Zhu, Nie, et al, ; Zhu, Quan, et al, ). In brief, maturated oocyte was enucleated by aspirating the first polar body plus a portion of the adjacent cytoplasm (presumably containing the metaphase II plate) using a sharp beveled glass pipette (WPI, Sarasota, FL) with a diameter of 20 ~ 25 µm.…”

“…For generation of cloned pigs, a surgical ET was performed as described previously (Nie et al, ; Zhu, Quan, et al, ). In brief, two to three hundreds of cloned embryos were cultured for 1 ~ 2 days in vitro after activation and then surgically transferred into the oviductal ampullary‐isthmic junction of the surrogates exhibiting natural oestrus (within 1 day of the onset of oestrus) using a sterility catheter (Covidien Inc, Mansfield, MA).…”

“…In addition, ear biopsies from transgenic‐cloned piglets and a WT control piglet were prepared for isolating the fibroblast cells according to described previously (Nie et al, ; Zhu, Nie, et al, ). The expression of eGFP in these in vitro cultured cells was examined under an Eclipse Ti fluorescence microscope (Nikon) equipped with filter set for eGFP.…”