The objective of this study was to obtain calves with an integrated human interferon alpha (IFNα) gene. The transgene (pbLGIFN-GFPBsd) was introduced by microinjection into one of the zygote’s pronuclei. The transgene (pbLGIFN-GFPBsd) contained the human interferon alpha (IFNα) gene, bovine lactoglobulin promoter (bLG), gfp (green fluorescence protein) and blasticidin resistance gene (bsd). After microinjection of 2107 zygotes, the proportion developing to the blastocyst stage (10.4%) was lower than for non-injected embryos (39.9%; P<0.01). Of the 2107 that were microinjected, 76 were GFP-positive blastocysts (3.6%). Fifty-seven GFP-positive blastocysts and 31 blastocysts from non-injected embryos were transferred singly to recipients. A lower rate of calving was observed after transfer of GFP-positive blastocysts (19.3%) as compared with noninjected embryos (48.4%; P<0.05). All newborn calves were healthy and of normal weight. PCR analyses indicated that none of the calves obtained after transfer of GFP-positive blastocysts carried the human interferon alpha gene.