2. Immunology Research Centre, St. Vincent's Hospital Melbourne, Melbourne, Victoria, Australia.
Several promoters from different origins have been used to drive transgene expression in pigs with variable results. In this study, we identified porcine ROSA26 promoter located on chromosome 13. Porcine ROSA26 promoter was cloned and its activity was examined using enhanced green fluorescence protein (EGFP) as a reporter. EGFP expression was detected in adult liver derived cells (LDC) by flow cytometry and confocal microscopy after transient transfection. Stable LDC expressing EGFP were established and used as nuclear donors for somatic cell nuclear transfer (SCNT). Two hundred thirty-three reconstructed embryos were transferred to two recipients. One pregnancy was established. A total of four fetuses were collected at day 30 of gestation, one normal and three resorbed. All fetuses had EGFP gene integration. Confocal microscope analysis showed that EGFP was expressed in fetal liver, heart, and isolated fetal fibroblasts. These results indicated that ROSA26 promoter is a ubiquitous promoter and can be utilized in pig transgenesis.
Keywords: ROSA26 locus, promoter, transgenesis, Enhanced Green Fluorescence Protein