cDNA library was constructed from the red Phalaenopsis petals by SMARTer technique.The F3′H unigene in Phalaenopsis was obtained through analyzing the EST sequences from library and it was analyzed using bioinformatics.At last,the expression patterns of F3′H in red and yellow Phalaenopsis petals were analyzed using real-time PCR.The results indicated that:(1)The recombinant percentage of the library was determined as 100.0% by random screening 24 clones.Among these clones,there were 22 inserts that were longer than 750 bp,up to 91.7%.(2)The amino acid sequence encoded by PhF3′H obtained from the library had a high identity with that of Dahlia pinnata (ADB_77826.1),Tricyrtis hirta (BAH_22519.1) and other ornamental plants.(3)The result of RT-PCR showed that PhF3′H expression abundance of red Phalaenopsis was about 19 times than that of the yellow Phalaenopsis,which proved that the gene plays an important role in the accumulation of anthocyanins in Phalaenopsis.In brief,the results will provide a theoretical basis to Phalaenopsis gene mining and color breeding.