Rubber seed oil (RSO) is a well-known non-edible oil currently not commercially used because no major application has been identified. Recently, studies of biodiesel produced from RSO have been reported. However, RSO biodiesel production potential and fossil diesel substitution potential have not been clarified. We report here estimates of the biodiesel production potential from rubber seed in natural-rubber-producing countries, with a combination of statistical data and original experimental data. The result shows that Indonesia has a RSO production potential of 889 098 tons/year as the highest country. In Nigeria, 15.1% of current fossil diesel consumption can potentially be replaced by biodiesel produced from an annual output of RSO locally. The usage of RSO for producing biodiesel is one option of reducing the biodiesel production dependency upon edible oil in these countries. The development of biodiesel production using RSO could play an important role in the natural rubber-producing countries, such as Nigeria, Indonesia, Thailand, Malaysia, Myanmar, and Ivory Coast.