The Pacific oyster, Crassostrea gigas, is a species in which true male and true female well as hermaphrodite individuals have been reported. Among the latter, most are sequential, with one or more sex changes throughout their live, while a few (<2%) are simultaneous hermaphrodites (SH) which may self-fertilize. Sex determinism and its functional bases remain unclear is this species. In this study, one SH oyster was found among a mature broodstock and used to produce a progeny by self-fertilization. Several thousand offspring were obtained for this family (SF1) but only 73 SF1 oysters survived following an episode of mass mortality at the spat stage. Six of them were conditioned for reproduction and three were found to be SH, and one produced a limited but viable progeny by self-fertilization (SF2). Oysters were sampled and genotyped with 226 SNP markers, confirming that the SF1 and the SF2 oysters were produced by self-fertilization over two successive generations. To our knowledge, it is the second study reporting the feasibility of production on an inbred line of Pacific oysters by self-fertilization using a simultaneous hermaphrodite as primary genitor, and the first study to generate two successive generations of selfing. Observed mean inbreeding coefficient of SF2 oysters produced by selfing for two generations was 0.61, ranging from 0.46 to 0.80. This coefficient was lower than the expected theoretical value (0.8125), suggesting that the most homozygous offspring died at early stages due to inbreeding depression. Our study demonstrates that inbred lines of Pacific oysters can be produced by self-fertilization but may exhibit higher heterozygosity than theoretically expected due to better survival of the most heterozygous offspring.