We isolated differentially expressed and dark-responsive genes during flower development and opening in petals of morning glory. Flower opening usually depends on petal expansion and is regulated by both genetic and environmental factors. Flower opening in morning glory (Ipomoea nil) is controlled by the dark/light regime just prior to opening. Opening was normal after 8- or 12-h dark periods but progressed very slowly after a 4-h dark period or in continuous light. Four genes (InXTH1-InXTH4) encoding xyloglucan endotransglucosylase/hydrolases (XTHs) and three genes (InEXPA1-InEXPA3) encoding alpha-expansins (EXPAs) were isolated. The expression patterns of InXTH2, InXTH3, and InXTH4 in petals were closely correlated with the rate of flower opening controlled by the length of the dark period prior to opening, but those of the EXPA genes were not. The expression pattern of InXTH1 gene was closely correlated with petal elongation. Suppression subtractive hybridization was used to isolate dark-responsive genes accompanying flower opening. The expressions of ten isolated genes were associated with the length of the dark period prior to flower opening. One gene was highly homologous to Arabidopsis pseudo-response regulator7, which is associated with the circadian clock and phytochrome signaling; another to Arabidopsis REVEILLE1, which affects the output of the circadian clock. Other genes were related to light responses, plant hormone effects and signal transduction. The possible roles of these genes in regulation of flower opening are discussed.