Abstract: As the main component of wood, lignin is usually polymerized from three monomers. During its biosynthesis, a total of ten enzyme families are involved in the conversion of phenylalanine to monomeric lignin, of which C3H is the key control enzymes in the hydroxylation process of p-coumaroyl CoA to caffeoyl CoA and in the formation of G/S monomer. Exploring the expression pattern of PagC3H3 gene is of great significance for further understanding of the function of this gene. In this study, the tissue-specific expression of PagC3H3 gene was analyzed by quantitative PCR; the promoter sequence of PagC3H3 with a length of 2 035 bp was cloned, and it was predicted to contain multiple cis-acting elements. At the same time, the promoter sequence of PagC3H3 was constructed to construct a plant expression vector pBI121-PagC3H3pro::GUS for transient transformation of Arabidopsis thaliana. The expression of PagC3H3 gene was higher in the roots, middle stem segments and basal stem segments of Populus alba×P. grandulos; transient transformation of A. thaliana showed that GUS activity was stronger in hypocotyls and roots, suggesting that PagC3H3 gene plays a role in lignin synthesis.