To investigate the function of GsF3′5′H and GsFNS in Gentiana sinoornata, with blue corolla during 5 different flowering stages (H₁-H₅) as the test material, we cloned the fulllength cDNA by using RTPCR techniques. The results showed that: (1) GsF3′5′H and GsFNS contain 1 560 bp and 1 590 bp open reading frame (OFR), which encode 520 and 529 amino acids, respectively. (2) Structural analysis revealed that both GsF3′5′H and GsFNS had typical conserved domains of F3′5′H and FNSⅡ. (3) Phylogenetic analysis indicated that Gentiana triflora was the closest relative species between GsF3′5′H and GsFNS. (4) qRTPCR showed that GsF3′5′H and GsFNS were expressed in roots, stems, leaves and corolla, and the expression level of GsF3′5′H was the highest in corolla H₃ stage, the expression level of GsFNS was the highest in roots, which followed by the corolla H₄ stage. It showed that the copigmentation effect of delphinin and flavone expressed by GsF3′5′H and GsFNS may make the corolla of Gentiana sinoornata more stable and bluer, which providing important genetic resources for the molecular breeding of blue flowers.