We investigated two putative gap genes encoding glyceraldehyde 3-phosphate dehydrogenase (GAPDH) in Giardia lamblia. While the expression of the gap2 gene was induced during encystation, gap1 was constitutively expressed. The induction of GAP2 protein during encystation was confirmed using GAP2-specific polyclonal antibodies. Complementation tests using a gap-deficient Escherichia coli showed that only the cell containing the gap1 gene, but not that with the gap2 gene, was able to restore GAPDH activity and grow in the presence of glucose as a sole carbon source. Levels of GAPDH activity were found to be constant in G. lamblia extracts at various times of encystation. These results suggest that a GAP1 protein functions as a major glycolytic enzyme, and that gap2 does not seem to encode a protein with GAPDH activity.