There have been many studies on biological hydrogen production thanks to its eco-friendly properties and low cost of facilities as hydrogen takes increasing interest as substitutive energy source. Up to now, most of researches focused on using hydrogen-producing microorganisms screened and isolated from nature. These microbes include algae, cyanobacteria, and anaerobic bacteria that produce hydrogen by degrading organic compounds. Biohydrogen production using them has many limitations because hydrogen production is only possible under anaerobic condition and cell growth rate is low compared to facultative chemotrophs. We are investigating on development of recombinant system by expression of hydrogenase capable of producing hydrogen which are isolated from various microorganisms. To do so, efficient and high throughput screening method for hydrogenase is necessary. We are doing researches on obtaining oxygen-tolerant hydrogenase based on these techniques and making recombinant E. coli system by expression of hydrogenase. We are also performing investigation on photo-conversion effects on biohydrogen production in recombinant E. coli .