Laparoscopic transfer of in vitro fertilized gene-edited embryos in pigs.

Laparoscopic embryo transfer in pigs has been explored as a minimally invasive alternative to conventional laparotomic methods, which often lead to surgical risks such as adhesions, prolonged recovery, and postoperative complications. This study is the first to report the generation of genetically modified pigs via embryo transfer using a laparoscopic approach. Gene-edited embryos were prepared by delivering the CRISPR/Cas9 system via electroporation to target the Coagulation Factor VIII, WDR45, and IL2RG genes and then transferred into the oviducts of five recipient gilts using a laparoscopic technique. Embryos putatively carrying modifications in the Coagulation Factor VIII and WDR45 genes were each transferred into a single recipient gilt, and both recipients became pregnant. In contrast, embryos putatively carrying a modification in the IL2RG gene were transferred into three recipient gilts, of which two became pregnant. In total, four recipient gilts were carried to full term, resulting in the birth of 9 live piglets. Genotyping confirmed that all of the offspring carried the genetic modifications, demonstrating this method may have potential for producing genetically modified pigs. The successful production of live gene-edited offspring demonstrates the feasibility of laparoscopic embryo transfer for genetically modified embryos. Although further research, including a large number of recipient pigs, as well as direct comparisons with conventional laparotomy, is needed to evaluate embryo survival, implantation efficiency, and animal welfare, laparoscopic embryo transfer represents a feasible approach for generating genetically modified pigs.