Animals as a source of organs and tissues for xenotransplantation could become a backup solution for the growing shortage of human donors. The presence of human xenoreactive anti- bodies directed against Galα1,3Gal antigens on the cell surface of a pig donor triggers the activa- tion of the complement leading to a hyperacute reaction. The development of genetic engineer- ing techniques has enabled the modification of genomes by knocking in and/or knocking out genes. In this paper, we report the generation of modified pigs with ZFN mediated disruption of the GGTA1 gene encoding the enzyme responsible for synthesis of Galα1,3Gal antigens. ZFN plasmids designed to target the exon 9 region of the pig GGTA1 gene encoding the catalytic domain were injected into the pronuclei of fertilized egg cells. Among 107 piglets of the F0 gene- ration analyzed, one female with 9-nt deletion in exon 9 of the GGTA1 gene was found. 13 of 33 piglets of the F1 generation represented the +/- GGTA1 genotype and 2 of 13 F2 piglets repre- sented the -/- GGTA1 genotype. No changes in the animals’ behavior, phenotype or karyotype were observed. Analysis confirmed heredity of the trait in all animals. A complex functional analysis of the modified animals, including flow cytometry, human serum cytotoxicity test and immunohistochemical detection, was performed to estimate the phenotype effect of genetic modification and this indicated an efficient GGTA1 knock-out in modified pigs.

The aim of the study was to develop a method of laparoscopic embryo transfer in pigs and to compare different variants of this method. Two catheter diameters (1.6 mm and 1.0 mm), the method and site of embryo deposition (oviduct or uterus), the embryo development stage (2 – 4 cell or blastocyst), the method for oviduct or uterus stabilization, the potential for cryopreserved embryo transfer, the developmental potential of the embryos after transfer to the oviduct, patomorphology of the oviduct after transfer and possible clinical complications were taken into consideration. Two studies compared two variants of transfer to the uterus, and five variants of transfer to the fallopian tube. The transfer of embryos by the infundibulum may be of limited use due to handling problems and very low efficiency (pregnancy was not achieved). Very low efficiency was shown after transfer of vitrified embryos. Transfer to the fallopian tube by puncture of the fallopian tube, regardless of the developmental stage of the embryo, is the recommended method of embryo transfer. The histopathological examination of the fallopian tube revealed possible changes within the puncture site. The numerous clinical complications observed did not affect the effectiveness of the method.