This study was conducted to evaluate developmental gene expression of lactoferrin and effect of supplementary iron on gene expression of lactoferrin in mice mammary gland using semi-quantitative RT-PCR analysis. In experiment 1, total twelve female ICR ( Institute of Cancer Research ) mice of four groups, each group of three mice at day 1, 9, 17 and 25 of lactation were used to determine effect of different lactating stages on mRNA expression of lactoferrin. In experiment 2, six groups mice (total twenty-four female mice at day 12 after mating) feeding purified diets (without iron or supplement iron) were assigned to two treatments (control and treatment). The feeding experimental period lasted 35 days. During feeding experiment, six mice with three animals each group were chosen at day 1, 9, 17 and 25 of lactation to determine the effect of iron on lactoferrin mRNA expression of mice at different lactating stages. The current results of experiment 1 showed that lactoferrin mRNA had strong expression at day 1 of lactation, and decreased gradually at day 9 and day 17 of lactation and then increased again markedly at day 25 of lactation. These results implicated that the expression of lactoferrin in mammary gland at different lactating stages is consistent with the changes of lactoferrin concentrations in milk. Iron significantly increased lactoferrin mRNA expression at day 1 and day 25 of lactation. Iron also increased lactoferrin gene expression at day 9 and day 17 of lactation, but statistical result was not significant. These findings raised the possibility that iron supplementation may play a role in regulation of lactoferrin levels in vivo.